Method Comparison Using 2 Point-of-Care Meters and a Reference Analyzer for Measuring Blood Triglycerides in Psittacine Birds.

Female reproductive disorders, such as chronic egg laying, are common in captive psittacine birds. While a disease diagnosis related to reproductive disorders can often be accomplished by physical examination and diagnostic imaging, monitoring of the response to environmental modification and medical treatment is more challenging. Monitoring ideally would involve measurement of luteinizing hormone or estrogen to assess ovarian activity. However, the amount of blood required for hormone analysis is greater than the small sample size that one can collect from these birds. Additionally, the lack of reference intervals limits their use as a diagnostic tool. Because plasma triglyceride increases during sustained estrogen release from the ovary, it may be used as an alternative method for assessing ovarian activity in birds. Point-of-care (POC) analyzers for measuring lipids in human plasma use very small sample volumes and have been used for measuring triglycerides in animals, including chickens. The authors therefore performed a method comparison study with 2 POC analyzers and a reference analyzer and plasma and whole blood from psittacine birds to determine whether these meters are suitable for triglyceride measurement in a known population of psittacine birds. Correlation, Deming regression, and Bland-Altman analyses were used to assess performance, and the total observed error for each meter relative to the reference analyzer was calculated. One of the meters exhibited fair performance and, with species-specific reference intervals, is likely to be clinically useful for triglyceride measurement in psittacine birds. The other meter demonstrated poor performance with unacceptable error, and its use for this purpose is strongly discouraged.

Can the intake of antiparasitic secondary metabolites explain the low prevalence of hemoparasites among wild Psittaciformes?

BACKGROUND: Parasites can exert selection pressure on their hosts through effects on survival, on reproductive success, on sexually selected ornament, with important ecological and evolutionary consequences, such as changes in population viability. Consequently, hemoparasites have become the focus of recent avian studies. Infection varies significantly among taxa. Various factors might explain the differences in infection among taxa, including habitat, climate, host density, the presence of vectors, life history and immune defence. Feeding behaviour can also be relevant both through increased exposure to vectors and consumption of secondary metabolites with preventative or therapeutic effects that can reduce parasite load. However, the latter has been little investigated. Psittaciformes (parrots and cockatoos) are a good model to investigate these topics, as they are known to use biological control against ectoparasites and to feed on toxic food. We investigated the presence of avian malaria parasites (Plasmodium), intracellular haemosporidians (Haemoproteus, Leucocytozoon), unicellular flagellate protozoans (Trypanosoma) and microfilariae in 19 Psittaciformes species from a range of habitats in the Indo-Malayan, Australasian and Neotropical regions. We gathered additional data on hemoparasites in wild Psittaciformes from the literature. We considered factors that may control the presence of hemoparasites in the Psittaciformes, compiling information on diet, habitat, and climate. Furthermore, we investigated the role of diet in providing antiparasitic secondary metabolites that could be used as self-medication to reduce parasite load. RESULTS: We found hemoparasites in only two of 19 species sampled. Among them, all species that consume at least one food item known for its secondary metabolites with antimalarial, trypanocidal or general antiparasitic properties, were free from hemoparasites. In contrast, the infected parrots do not consume food items with antimalarial or even general antiparasitic properties. We found that the two infected species in this study consumed omnivorous diets. When we combined our data with data from studies previously investigating blood parasites in wild parrots, the positive relationship between omnivorous diets and hemoparasite infestation was confirmed. Individuals from open habitats were less infected than those from forests. CONCLUSIONS: The consumption of food items known for their secondary metabolites with antimalarial, trypanocidal or general antiparasitic properties, as well as the higher proportion of infected species among omnivorous parrots, could explain the low prevalence of hemoparasites reported in many vertebrates.

Concentrations of calcium and 25-hydroxycholecalciferol (vitamin D3) in plasma of wild kakapo (Strigops habroptilus) living on two islands in New Zealand.

AIMS This preliminary study had the objectives of describing the concentrations of ionised calcium and 25-hydroxycholecalciferol (25(OH)D3) in the blood of wild kakapo (Strigops habroptilus) living on two islands in New Zealand, and to determine the effects of supplementary feeding on these blood parameters. METHODS Blood samples were obtained from 33 kakapo living on two offshore islands during routine health checks in 2015. Birds on Hauturu were sampled in May (n=5) and birds on Whenua Hou were sampled in July (n=15) and November (n=26). Of the birds sampled on Whenua Hou in November, 15 received supplementary food prior to sampling. Samples were analysed for pH, and concentrations of ionised calcium, total calcium, phosphorous, total protein, albumin, globulin, uric acid and 25(OH)D3. RESULTS Concentrations of ionised calcium did not differ between unsupplemented birds on the two islands, nor between supplemented (median 1.17 (95% CI=1.12-1.20) mmol/L) and unsupplemented (median 1.09 (95% CI=1.08-1.14) mmol/L) birds sampled in November on Whenua Hou (p>0.05), and were comparable with published normal ranges for other psittacines. Concentrations of 25(OH)D3 did not differ between unsupplemented birds on the two islands (p>0.05), but were higher in supplemented (median 8.00 (95% CI=4.76-8.45) nmol/L) than unsupplemented (median 0.00 (95% CI=-0.16-0.48) nmol/L) birds on Whenua Hou (p<0.001). All values were much lower than published ranges for healthy psittacines. There was no difference between male and female birds on Whenua Hou for any parameter measured (p>0.05). CONCLUSIONS AND CLINICAL RELEVANCE The calcium status of the kakapo in this study was comparable to other wild psittacines, however concentrations of 25(OH)D3 were much lower. The concentrations of 25(OH)D3 may be within the normal range for the species, however further data are required to confirm this. The significant increase in concentrations of 25(OH)D3 in supplementary fed birds suggests that this food was providing more of the nutrient than the wild diet at that time of year, although the effects of this are unknown. Further investigation is required into the calcium and vitamin D3 status of kakapo, across a wider range of locations, seasons and ages. This would help define normal ranges for these parameters, allow interpretation in clinically abnormal individuals, and guide the refinement of supplementary foods. This information would, therefore, assist the future conservation management of this critically endangered species.

Blood Biochemical Values of Wild Scarlet Macaw ( Ara macao macao) Nestlings and Adults.

Blood biochemical values are useful as indicators of disease in veterinary practice and for health assessments of free-ranging animal populations. We analyzed blood biochemical values for free-living nestling and adult scarlet macaws ( Ara macao macao) in southeastern Peru with an Abaxis VetScan VS2 portable analyzer with Avian/Reptilian Profile Plus rotors. The resulting data were used to create provisional age-independent reference intervals for this instrument for wild scarlet macaw nestlings, calculate blood biochemical ranges for this instrument for wild adults, and examine age-related trends in nestlings. Levels of 11 parameters were studied: albumin, aspartate aminotransferase, calcium, creatine kinase, globulin, glucose, phosphorus, potassium, total protein, sodium, and uric acid. Bile acid levels were generally below the instrument detection level and were not evaluated. Most values and their trends with bird age were comparable to those observed in captive large macaw nestlings. Albumin, aspartate aminotransferase, globulin, glucose, total protein, sodium, and uric acid levels increased with nestling age to adulthood. Creatine kinase, phosphorus, and potassium concentrations decreased with age to adulthood. Calcium concentrations did not change between nestlings and adults. These changes in values with age are broadly in agreement with values in other avian species and likely reflect physiologic and developmental changes as nestlings mature.

Impact of Delayed Analysis in Avian Blood Biochemical Values Measured With the Abaxis VetScan VS2.

For biochemical analysis with a point-of-care biochemical analyzer, standard procedure is to analyze the sample as rapidly as possible (<1 hour) after venipuncture to minimize any changes in analyte concentrations that might occur over time. However, under some circumstances, such as when collecting blood at remote field sites, a longer delay may be unavoidable. This study evaluates the effect of delayed analysis time under unrefrigerated conditions on avian (psittacine) biochemical analyte concentrations obtained with the VetScan VS2 using Avian/Reptilian Profile Plus rotors. Venipuncture was performed on a group of 36 psittacine birds as part of routine health checks in a research aviary (Texas A&M University, College of Veterinary Medicine, College Station, TX, USA). Whole blood was kept at room temperature and analyzed at 4 time intervals: <1, 3, 6, and 24 hours after venipuncture. At 3 hours or less after collection, most biochemical parameters changed by <2%, with the exception of phosphorus (decrease of about -9%). Major increases by 24 hours after collection were observed in phosphorus (+67%) and potassium (+103%) concentrations, whereas aspartate aminotransferase (AST), uric acid, glucose, and sodium concentrations also showed statistically significant changes. Our results suggest that accurate information from analyses using the VetScan VS2 may be obtained for up to 3 hours after venipuncture without refrigeration, but researchers and clinicians do need to exercise care when interpreting blood chemistry analyte concentrations obtained after multihour delays between venipuncture and sample analysis.

Plasma butyrylcholinesterase concentrations in psittacine birds: reference values, factors of variation, and association with feather-damaging behavior.

Butyrylcholinesterase is a glycoprotein enzyme used in the diagnosis of toxicosis by cholinesterase-inhibitor agents like organophosphates and carbamates. In animals, butyrylcholinesterase concentrations have been shown to vary depending on numerous factors such as age, sex, diet, and season of sampling. To establish reference values of plasma butyrylcholinesterase concentrations in common psittacine species, plasma butyrylcholinesterase concentrations were measured in 1942 companion psittacine birds. The birds were classified by age, sex, season, health status, and the presence of feather-damaging behavior. A significant difference was observed among species, with eclectus parrots (Eclectus roratus) having the lowest and African grey parrots (Psittacus erithacus) having the highest reference values. Plasma butyrylcholinesterase concentrations varied by age, health status, and season but not by sex. Concentrations were significantly higher during autumn and spring than during winter and summer, and significantly lower in healthy birds than in sick birds. No significant association between butyrylcholinesterase concentrations and feather-damaging behavior could be established except in lovebirds (Agapornis species). Further research is needed to better understand the effect of nutritional and hormonal factors on butyrylcholinesterase concentrations in psittacine birds and its possible effect on bird cognition.

Use of an indirect sampling method to produce reference intervals for hematologic and biochemical analyses in psittaciform species.

As with other animal species, comprehensive reference intervals (RI) for psittaciform species are rare and plagued by common issues, including sparse information regarding methods used to analyze specimens, low sample sizes, and improper statistical analyses. The purpose of this study was to examine the use of an indirect sampling method of RI generation from several years of data collected from specimens of multiple psittaciform species submitted to a veterinary diagnostic laboratory. These data were unselected for health status. A previously published method for indirect RI generation was applied to data collected for routine hematologic and biochemical analyses. Seven species groups were examined, and sample size ranged from 346 to 2358. Results showed that RI varied by species and appeared to represent a broader range than expected compared with other RI and traditional clinical expectations for core health assessments, such as total white blood cell count and white blood cell differential results. Some biochemical results reflected more narrow ranges, and a few were consistent with other published ranges. The intervals were likely influenced by changes related to stress and underlying disease. The results of the current study reflect the imprecision of this method related to data obtained from the population served by this laboratory. Overall, this method is not suitable for the production of comprehensive RI, although it may provide rough estimates for some limited analyses until traditional RI can be generated.

Complete blood cell count in psittaciformes by using high-throughput image cytometry: a pilot study.

The avian hemogram is usually performed in veterinary diagnostic laboratories by using manual cell counting techniques and differential counts determined by light microscopy. There is no standard automated technique for avian blood cell count and differentiation to date. These shortcomings in birds are primarily because erythrocytes and thrombocytes are nucleated, which precludes the use of automated analyzers programmed to perform mammal complete blood cell counts. In addition, there is no standard avian antibody panel, which would allow cell differentiation by immunophenotyping across all commonly seen bird species. We report an alternative hematologic approach for quantification and differentiation of avian blood cells by using high-throughput image cytometry on blood smears in psittacine bird species. A pilot study was designed with 70 blood smears of different psittacine bird species stained with a Wright-Giemsa stain. The slides were scanned at 0.23 microm/pixel. The open-source softwares CellProfiler and CellProfiler Analyst were used for analyzing and sorting each cell by image cytometry. A "pipeline" was constructed in the CellProfiler by using different modules to identify and export hundreds of measures per cell for shape, intensity, and texture. Rules for classifying the different blood cell phenotypes were then determined based on these measurements by iterative feedback and machine learning by using CellProfiler Analyst. Although this approach shows promises, avian Leukopet results could not be duplicated when using this technique as is. Further studies and more standardized prospective investigations may be needed to refine the "pipeline" strategy and the machine learning algorithm.

Perfil hematológico, bioquímico sérico, proteína C reativa e cortisol de ararajubas (Guaroba guarouba) mantidas em cativeiro / Blood profile, serum biochemistry, C-reactive protein and cortisol in golden conures (Guaroba guarouba) in captivity

Objetivou-se determinar os valores hematológicos, bioquímicos séricos, proteína C reativa e cortisol de 14 ararajubas (Guaroba guarouba) clinicamente saudáveis mantidas em cativeiro no zoológico do Parque Estadual de Dois Irmãos Recife/PE. Amostras de sangue foram obtidas da veia jugular com volume médio de 0,8ml, fracionando em duas porções, a primeira depositada em tubo MiniCollect®CE contendo EDTA e a outra tubo com gel separador, para a separação do soro sanguíneo. Os indicadores bioquímicos e PCR foram determinados através do analisador ARCHITECT c8000. O cortisol foi analisado no Cobas E411 da Roche. Para a determinação dos números de eritrócitos e leucócitos, foi utilizada a metodologia da contagem em câmara de Neubauer. O hematócrito foi avaliado pelo método do microhematócrito e a hemoglobina pelo método da cianometahemoglobulina. Para a contagem diferencial de leucócitos, foi utilizada a técnica de Shilling. Os índices hematimétricos (VCM, HCM E CHCM) foram determinados com os valores encontrados na série eritrocítica. Os dados foram caracterizados por dispersão de freqüências, utilizando-se as seguintes medidas de tendência central: Média, desvio-padrão, mediana e percentil de 25 e percentil 75. A determinação de alguns parâmetros como ferro, triglicerídeos, PCR e cortisol foram identificados como sendo pioneiramente referenciados nesta espécie de ave. Os dados são apresentados como sendo de referência para a ararajuba (Guaroba guarouba) criada em cativeiro em condições similares de manejo e higidez e ainda pode contribuir para os trabalhos de conservação ex situ desta espécie.

The aim of this study was to determine the hematological, serum biochemical, C-reactive protein and cortisol 14 clinically healthy golden conures (Guaroba guarouba) kept in captivity at the Zoo of Parque Estadual de Dois Irmãos Recife/PE, Brazil. Blood samples were collected from the jugular vein with an average of 0.8mL and divided into two parts, the first deposited in MiniCollect®EC and the other containing EDTA tube with gel separator for separating blood serum. Biochemical indicators and PCR was determined using the ARCHITECT analyzer c8000. Cortisol was analyzed on the Roche Cobas E411. To determine the number of erythrocytes and leukocytes the methodology of counting in a Neubauer chamber was used. Hematocrit was assessed by using the microhematocrit and hemoglobin for the method of cianometahemoglobulina. For the differential count the Shilling technique was used. The RBC indices (MCV, MCH and MCHC) were determined with values found in the erythrocytic series. The data were characterized by dispersion of frequencies, using the following measures of central tendency: mean, standard deviation, median and percentile 25 and 75 percentile. Parameters such as iron, triglycerides, CRP, and cortisol were identified and referenced in this pioneering species of bird. Data are presented as reference for Guaroba guarouba raised in captivity under similar conditions of management and healthiness and can still contribute to the work of ex situ conservation of this species.

Overview of psittacine blood analysis and comparative retrospective study of clinical diagnosis, hematology and blood chemistry in selected psittacine species.

Part 1 presents a comprehensive overview of the differences between mammals on sampling methods, processing, testing, and interpretation of data, with special attention to the meaning of pathologic and normal, and the differences among species and diseases. Part 2 critically analyzes 150 different hematologic and biochemical profiles obtained over 5 years from 150 psittacine birds belonging to 29 different species, brought to our clinic and/or examined in other locations, with confirmed or strongly suspected diseases. The results are grouped according to the various pathologies observed with respect to species and the degree of variation from the reference range of each species.

Pharmacokinetics after intravenous administration of flunixin meglumine in budgerigars (Melopsittacus undulatus) and Patagonian conures (Cyanoliseus patagonus).

OBJECTIVE: To investigate the disposition kinetics of flunixin meglumine when administered IV to budgerigars (Melopsittacus undulatus) and Patagonian conures (Cyanoliseus patagonus). DESIGN: Prospective cohort study. ANIMALS: 8 adult Patagonian conures and 24 adult budgerigars. PROCEDURES: Injectable flunixin meglumine (50 mg/mL) was diluted to 10 and 1. 0 mg/mL and administered IV at a dose of 5.0 mg/kg (2.3 mg/lb) to Patagonian conures and budgerigars, respectively. RESULTS: In budgerigars, the elimination half-life was 0.72 hours and the mean residence time was 0.73 hours. In Patagonian conures, the elimination half-life was 0.91 hours and the mean residence time was 1.20 hours. The concentration of flunixin was below the assay's limit of quantification (0.5 µg/mL) at 3 and 6 hours in budgerigars and Patagonian conures, respectively. A single budgerigar developed adverse effects (lethargy and signs of depression) for approximately 15 minutes following drug administration. CONCLUSIONS AND CLINICAL RELEVANCE: The half-life of flunixin in Patagonian conures and budgerigars was short following IV administration; however, results of this study suggested that IV administration of injectable flunixin meglumine at 5.0 mg/kg resulted in plasma concentrations that could potentially be anti-inflammatory and analgesic in budgerigars and Patagonian conures.

Use of refractometry for determination of psittacine plasma protein concentration.

BACKGROUND: Previous studies have demonstrated both poor and good correlation of total protein concentrations in various avian species using refractometry and biuret methodologies. OBJECTIVES: The purpose of the current study was to compare these 2 techniques of total protein determination using plasma samples from several psittacine species and to determine the effect of cholesterol and other solutes on refractometry results. METHODS: Total protein concentration in heparinized plasma samples without visible lipemia was analyzed by refractometry and an automated biuret method on a dry reagent analyzer (Ortho 250). Cholesterol, glucose, and uric acid concentrations were measured using the same analyzer. Results were compared using Deming regression analysis, Bland-Altman bias plots, and Spearman's rank correlation. RESULTS: Correlation coefficients (r) for total protein results by refractometry and biuret methods were 0.49 in African grey parrots (n=28), 0.77 in Amazon parrots (20), 0.57 in cockatiels (20), 0.73 in cockatoos (36), 0.86 in conures (20), and 0.93 in macaws (38) (P< or =.01). Cholesterol concentration, but not glucose or uric acid concentrations, was significantly correlated with total protein concentration obtained by refractometry in Amazon parrots, conures, and macaws (n=25 each, P<.05), and trended towards significance in African grey parrots and cockatoos (P=.06). CONCLUSIONS: Refractometry can be used to accurately measure total protein concentration in nonlipemic plasma samples from some psittacine species. Method and species-specific reference intervals should be used in the interpretation of total protein values.

Parâmetros de bioquímica sérica de machos, fêmeas e filhotes de Araras canindé (Ara ararauna) saudáveis mantidas em cativeiro comercial / Serum biochemical parameters of healthy male, female and young blue-and-yellow macaws (Ara ararauna) bred in captivity

O Brasil possui um considerável número de espécies de psitacídeos catalogados, perfazendo cerca de 80 espécies, sendo que as Araras canindé (Ara ararauna), uma das maiores representantes dessa ordem, podem ser encontradas em florestas nas diversas regiões brasileiras. O Instituto Brasileiro do Meio Ambiente e dos Recursos Naturais Renováveis (IBAMA) normalizou a comercialização de animais da fauna silvestre provenientes de criadouros e conseqüentemente ocorreu um aumento do número destes animais como de estimação. Atualmente, há pouco conhecimento sobre os parâmetros clínicos e laboratoriais de espécies silvestres em cativeiro. O presente trabalho teve como objetivo determinar parâmetros de bioquímica sangüínea de Araras canindé (Ara ararauna) saudáveis de sexo e faixa etária distintas mantidas em um criatório comercial com alimentação e manejo controlados e padronizados. Foram colhidas amostras de sangue de 35 araras canindé (11 filhotes e 24 adultos) e remetidas ao Laboratório de Análises Clínicas Veterinárias da Universidade de Passo Fundo (UPF) para determinação dos seguintes indicadores bioquímicos: ácido úrico (AU), albumina (ALB), aspartato aminotransferase (AST), cálcio (Ca), colesterol (Col), creatina quinase (CK), fosfatase alcalina (FA), fósforo inorgânico (Pi), frutosamina (Fru), gama glutamil transferase (GGT), proteínas totais (PT) e uréia (UR). Entre as aves adultas, foram encontradas diferenças significativas nos valores de CK (superior nos machos), Ca e AU (superiores nas fêmeas). Entre aves adultas e filhotes foram constatadas diferenças significativas nos valores de AU, Ca, ALB, COL, FA, Pi e FRU. A maioria das alterações detectadas pode ser relacionada à diferença na dieta fornecida e à condição fisiológica de cada categoria de aves. Os dados obtidos podem ser utilizados como parâmetros de referência para as araras canindé brasileiras.

Several psittacines have been cataloged in Brazil, totaling nearly 80 species. The blue-and-yellow macaw (Ara ararauna), the most representative species of the Psittaciformes order, lives in the forest areas of different Brazilian regions. IBAMA, Brazilian environmental protection agency, has authorized the trade of wild animals raised in breeding facilities; consequently, their use as household pets has increased ever since. Currently, too little is known about clinical and laboratory parameters of wild species bred in captivity. The aim of the present study was to determine the serum biochemical parameters of healthy blue-and-yellow macaws (Ara ararauna) of different sexes and ages reared in a commercial breeding facility under controlled and standardized feeding and management practices. Blood samples were collected from 35 blue-and-yellow macaws (11 fledglings and 24 adults) and sent to the Laboratory of Veterinary Clinical Pathology of Universidade de Passo Fundo (UPF) for measurement of the following biochemical parameters: uric acid (UA), albumin (Alb), aspartate aminotransferase (AST), calcium (Ca), cholesterol (Chol), creatine kinase (CK), alkaline phosphatase (AP), inorganic phosphorus (iP), fructosamine (Fru), gamma-glutamyl transferase (GGT), total protein (TP) and urea nitrogen (Ur). There were significant differences among adult macaws with regard to CK levels (higher in males) and to Ca and UA levels (higher in females). Differences were noted among adults and fledglings with respect to UA, Ca, Alb, Chol, AP, iP and Fru. Most of the findings can be ascribed to the different feeding regimens and ages of the birds analyzed. The data obtained herein can be used as reference parameters for Brazilian blue-and-yellow macaws.

Assessment of a point-of-care biochemical analyzer and comparison with a commercial laboratory for the measurement of total protein and albumin concentrations in psittacines.

OBJECTIVE: To determine agreement for total protein (TP) and albumin concentrations measured by a point-of-care biochemical analyzer in heparinized whole blood and plasma samples obtained from psittacines and compare results with those from a commercial laboratory. SAMPLE POPULATION: Hematologic samples from 92 healthy birds. PROCEDURES: Duplicate samples of heparinized whole blood and plasma were obtained. A point-of-care biochemical analyzer was used to determine TP and albumin concentrations. To assess precision, intraclass correlation coefficient (r(i)) and Bland-Altman measures of agreement were used. These results were compared by use of Bland-Altman plots with those obtained from a commercial laboratory that used a biuret method for TP concentration and electrophoresis for albumin concentration. RESULTS: For the analyzer, there was excellent agreement (r(i) = 0.91) between heparinized whole blood and plasma samples for TP and albumin concentrations. Relative error was 0.9% for TP and 0.7% for albumin. Analyzer results correlated well with commercial laboratory results, with a downward bias of 0.6 for TP and 0.3 for albumin. CONCLUSIONS AND CLINICAL RELEVANCE: The analyzer had excellent precision for analysis of heparinized whole blood or plasma samples for TP or albumin concentrations; analyzer values had good agreement with those from a commercial laboratory. The analyzer could be a valid method to measure plasma TP concentrations and provide point-of-care testing in apparently healthy parrots. Biochemical analyzer results for plasma albumin concentration were not validated by results from a commercial laboratory, so conclusions cannot be drawn regarding use of the analyzer in measurement of albumin concentrations in psittacines.

Protein electrophoresis of psittacine plasma.

BACKGROUND: Although protein electrophoresis (EPH) has been widely applied in human and veterinary medicine, it has only recently been implemented in the analysis of avian samples. OBJECTIVE: The purpose of this study was to examine the application of protein EPH to the analysis of psittacine plasma samples. Our goals were to describe protein fraction mobility, establish reference intervals for some common species, determine the coefficient of variation (CV) of the chosen method, and examine the effects of sample handling and sample condition. METHODS: Heparinized plasma samples from several common psittacine species (minimum sample size 50 each) were examined using the Beckman Paragon system and SPEP-II gels. Total protein was measured by refractometry. Reference intervals (95%) were calculated by the rank methods. RESULTS: Fraction migration patterns were found to vary among common psittacine species. Day-to-day CV for the EPH fractions ranged from 2.2% to 10.5%; within-run CV ranged from 4.8% to 10.8%; and total CV ranged from 3.2% to 14.8%. The highest CV was noted for the poorly defined alpha-globulin fraction. Prolonged refrigeration, repeated freeze-thawing, hemolysis, and lipemia altered the results. CONCLUSIONS: Protein fractions from psittacine species were variable in terms of migration pattern and protein concentration, which necessitates the use of species-specific reference intervals. Avian protein electrophoretic patterns and values should be interpreted based on knowledge of the CV associated with the technique as well as on the effects of sample handling and condition.

Pharmacokinetics of marbofloxacin in blue and gold macaws (Ara ararauna).

OBJECTIVE: To determine the pharmacokinetics of marbofloxacin after single IV and orally administered doses in blue and gold macaws. ANIMALS: 10 healthy blue and gold macaws. PROCEDURES: In a crossover study, marbofloxacin (2.5 mg/kg) was administered orally (via crop gavage) to 5 birds and IV to 5 birds. Blood samples were obtained at 0, 0.5, 1, 3, 6, 12, 24, 48, 72, and 96 hours after marbofloxacin administration. After a 4-week washout period, the study was repeated, with the first 5 birds receiving the dose IV and the second 5 birds receiving the dose orally. Serum marbofloxacin concentrations were quantitated by use of a validated liquid chromatography-mass spectrometry assay. RESULTS: After oral administration, mean +/- SD area under the curve was 7.94 +/- 2.08 microg.h/mL, maximum plasma concentration was 1.08 +/- 0.316 microg/mL, and bioavailability was 90.0 +/- 31%. After IV administration of marbofloxacin, the apparent volume of distribution was 1.3 +/- 0.32 L/kg, plasma clearance was 0.29 +/- 0.078 L/h/kg, area under the curve was 9.41 +/- 2.84 microg.h/mL, and the harmonic mean terminal half-life was 4.3 hours. CONCLUSIONS AND CLINICAL RELEVANCE: Single IV and orally administered doses of marbofloxacin were well tolerated by blue and gold macaws. The orally administered dose was well absorbed. Administration of marbofloxacin at a dosage of 2.5 mg/kg, PO, every 24 hours may be appropriate to control bacterial infections susceptible to marbofloxacin in this species.

Psittacine plasma concentrations of elements: daily fluctuations and clinical implications.

During the past 2 decades, the potential for excessive exposure of pet birds to zinc has become a concern for many pet bird owners. Ideally, avian zinc toxicosis is diagnosed on the basis of history of exposure to zinc, radiographic evidence of ingested metal, occurrence of melena, detection of an elevated plasma zinc concentration, and response to treatment. However, most pet birds suspected of having zinc toxicosis present with vague signs and lack of radiographic evidence; therefore, the diagnosis relies on the presence of an elevated plasma zinc concentration. A question was, is there a significant diurnal variation in the zinc concentration in psittacine birds and could this be clinically relevant? Because studies in other species have shown that zinc is not the only element that shows a diurnal variation, the authors examined 13 other plasma elements including arsenic, cadmium, calcium, cobalt, copper, iodine, iron, magnesium, manganese, molybdenum, potassium, selenium, and total phosphorus. Fifteen adult psittacine birds housed in the same aviary were used in this study. Three blood samples, separated by 4 hours, were taken from the right jugular vein in each bird. All elements were measured in plasma. Zinc, copper, and molybdenum revealed diurnal fluctuations. The results of this study suggest that interpretation of clinical samples may be more complicated than previously believed. Furthermore, on the basis of the results of this study, it is possible that some avian reference ranges may need to be reexamined.

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.

Evidence of unique genotypes of beak and feather disease virus in southern Africa.

Psittacine beak and feather disease (PBFD), caused by Beak and feather disease virus (BFDV), is the most significant infectious disease in psittacines. PBFD is thought to have originated in Australia but is now found worldwide; in Africa, it threatens the survival of the indigenous endangered Cape parrot and the vulnerable black-cheeked lovebird. We investigated the genetic diversity of putative BFDVs from southern Africa. Feathers and heparinized blood samples were collected from 27 birds representing 9 psittacine species, all showing clinical signs of PBFD. DNA extracted from these samples was used for PCR amplification of the putative BFDV coat protein (CP) gene. The nucleotide sequences of the CP genes of 19 unique BFDV isolates were determined and compared with the 24 previously described sequences of BFDV isolates from Australasia and America. Phylogenetic analysis revealed eight BFDV lineages, with the southern African isolates representing at least three distinctly unique genotypes; 10 complete genome sequences were determined, representing at least one of every distinct lineage. The nucleotide diversity of the southern African isolates was calculated to be 6.4% and is comparable to that found in Australia and New Zealand. BFDVs in southern Africa have, however, diverged substantially from viruses found in other parts of the world, as the average distance between the southern African isolates and BFDV isolates from Australia ranged from 8.3 to 10.8%. In addition to point mutations, recombination was found to contribute substantially to the level of genetic variation among BFDVs, with evidence of recombination in all but one of the genomes analyzed.

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.