Atherosclerosis is a disease of elastic and muscular arteries characterized by vascular inflammation and buildup of cholesterol, fibrin, calcium, and cellular waste products within the intima of the vessel wall. The buildup results in plaque formation, vascular remodeling, luminal obstruction, alterations of blood flow, and decreased oxygen supply to organs. It is primarily a disease of the aorta and great vessels (large and medium-sized arteries) and is characterized by proliferative vascular smooth muscle cells, collagen deposits, and varying amounts of intracellular and extracellular lipids, primarily cholesterol and cholesterol esters that accumulate between the basement membrane and the vascular endothelial cells of the tunica intima.1 Clinical signs usually develop when blood flow is restricted because of an occluded vessel or when thrombi break free and occlude smaller vessels. In human patients, the clinical manifestations of atherosclerosis include coronary artery disease, stroke, abdominal aortic aneurysm, and peripheral vascular disease.
Atherosclerosis is best described as a chronic immunoinflammatory, fibroproliferative disease fueled by lipid deposition, and endothelial cells, leukocytes, and intimal smooth muscle cells are the major factors in the development of this disease. Vascular disease in avian species includes etiologies and lesions similar to those seen in mammals, and atherosclerosis is considered the most common vascular disease identified in birds.2 Arteriosclerotic plaques composed of fibrous tissue between the intima and internal elastic lamina are common findings in many species of birds and have been founds in chickens as young as 4 weeks of age. These plaques are seen grossly as pale areas of increased thickening of the intima and can extend as circumferential lesions resulting in severe luminal narrowing. In birds, chondroid or osseus metaplasia as well as mineralization can be seen in the lesions.3–6
Much is known about atherosclerosis in nonpsittacine species of birds, and numerous reports7–15 exist. Atherosclerosis has been reported as the most common pathological change observed in blood vessels of captive psittacine birds.16 Atherosclerosis in captive psittacine birds is a postmortem finding at necropsy rather than an antemortem diagnosis. Findings in a retrospective study17 on atherosclerosis in psittacine birds indicated that Amazon parrots are the most commonly affected species and that age may be a risk factor. Krautwald-Junghanns et al18 reported the presence of atherosclerosis in 14 of 107 (13%) psittacine birds as determined by necropsy examination of the heart, with increased occurrence in Amazon parrots as described previously by Johnson et al.17
Fricke et al6 evaluated the pathogenesis of atherosclerosis in birds via immunohistochemistry as a means to demonstrate lymphocytes, macrophages, smooth muscle cells, and chondroitin sulfate in plaques. Although the response to injury hypothesis is currently well accepted in the pathogenesis of atherosclerosis, results of that study6 suggest that this hypothesis seems inapplicable in parrots on the basis of the lack of lymphocytes and macrophages and the absence of invasion and proliferation of smooth muscle cells in each atherosclerotic stage. These authors also identified alterations of organs (ie, arterial thrombosis in the liver and lung, pulmonary congestion with fibrosis, and hepatic congestion with centrolobular necrosis) that significantly correlated with atherosclerosis of the aorta.6
Little is known about risk factors for the development of atherosclerosis in psittacine birds because most studies17,19–22 have investigated prevalence. Known risk factors in human patients include obesity, gender, age, family history, high serum cholesterol and triglyceride concentrations, diabetes mellitus, cigarette smoking, alcohol consumption, hypertension, hyperhomocystinemia, systemic inflammation, and certain infectious diseases, most notably Chlamydophila pneumoniae.23 This widespread respiratory pathogen has been shown in numerous studies24–28 to be strongly associated with coronary artery disease in humans as determined by findings on PCR assay, immunohistochemical evaluation, ELISA, and microbial growth in tissue culture.
In pet psittacine birds, suspected risk factors include high cholesterol and triglyceride concentrations, dietary practices, captive care, sex, age, species, obesity, social stress and inactivity (sedentary lifestyle), and infectious diseases, specifically Chlamydophila psittaci infection. Finlayson and Hirchinson29 experimentally induced severe atherosclerosis and hypercholesterolemia in budgerigars by feeding diets supplemented with 2% cholesterol. A review of cases from the Philadelphia Zoo showed that dietary modification from a seed-based to a complete pellet diet resulted in a decrease in the severity of atherosclerosis in the great vessels of parrots.30 One study19 showed that increased intake of α-linolenic acid may have a protective affect against the development of atherosclerosis in parrots.
Chlamydial organisms have been isolated from approximately 150 avian species and are commonly found in psittacine birds,31 especially cockatiels and budgerigars. Among caged, nonpsittacine birds, infection with Chlamydiaceae organisms occurs most frequently in pigeons and doves and is infrequently diagnosed in canaries and finches. Chlamydophila psittaci is a bacterium that has zoonotic potential. In humans, the resulting infection is referred to as psittacosis (also called parrot fever and ornithosis). The purpose of the study reported here was to determine whether, similar to the association with C pneumoniae in humans, infection with C psittaci as well as other specific factors was associated with atherosclerosis in psittacine birds.
Materials and Methods
Criteria for selection of cases—Psittacine birds with a histopathologic diagnosis of atherosclerosis made between January 1994 and October 2003 at The Animal Medical Center in New York City were identified by a review of necropsy reports from that period. Birds were excluded if a complete medical record was not available and if paraffin-embedded tissues were not available containing the aorta and great vessels of the heart. Psittacine birds necropsied during the same period that did not have atherosclerosis and had available medical records and paraffin-embedded sections of aorta or great vessels of the heart were eligible for selection as control birds. Control birds were selected by convenience from among the different species origin groups such that the distribution of origin groups was similar to that of the general population of birds that were seen at The Animal Medical Center between 1994 and 2003 on the basis of a computer review of electronic medical records (data not shown).
Procedures—Necropsy reports and medical records of psittacine birds meeting the inclusion criteria were reviewed, and information including signalment, weight, diet, relevant history, and all clinicopathologic test results was recorded. Birds were grouped by natural origin as New World (South American), African, or Australasian species, and the percentage of each group was determined. Birds were also grouped on the basis of clinical signs at the time of initial evaluation and assigned into the following 5 groups: neurologic, gastrointestinal, respiratory, reproductive, or nonspecific. Information on whether the birds were brought to the hospital on an emergency basis or for a scheduled appointment was also obtained. Psittacine birds without atherosclerosis were selected from all birds that underwent necropsy during the study period and were included as control birds. A computer review of medical record information on all psittacine birds examined at The Animal Medical Center during the study period was performed to determine the demographic population; the percentages of admitted species closely matched the distribution for control birds. Available tissue specimens from all birds were reviewed by a pathologist to confirm the presence (study birds) and absence (control birds) of atherosclerosis. Paraffin-embedded sections of the great vessels containing the atherosclerotic plaques of the study birds and similar sections of the great vessels from control birds were prepared and reviewed. Slides were sent to the University of Georgia for immunohistochemical staining that was specific for C psittaci antigen.a
Immunohistochemical staining for C psittaci antigen—Staining via immunohistochemistry for detection of C psittaci antigen was performed with the avidin-biotin complex technique. Replicate paraffin-embedded tissue sections were dewaxed in limonene and rehydrated through graded alcohols to aqueous buffer. Endogenous peroxidase activity was quenched by treatment with 3% hydrogen peroxide solution. Antigenic sites were subsequently unmasked by treatment with steam and citrate buffer. Nonspecific binding of the primary antibody was minimized by treatment with a blocking reagent. A primary mouse anti–C psittaci monoclonal antibodyb was applied at a 1:1,500 dilution. Following incubation and washing, a secondary goat anti-rabbit, biotinylated antibody was applied. The tissue sections were incubated and washed again. The avidin-biotin complex reagent was applied and incubated. The avidin-biotin complex reagent consists of streptavidin that contains 4 binding sites for biotin. Three of these binding sites are occupied by biotin that is conjugated to alkaline phosphatase. The remaining binding site attaches to the biotinylated secondary antibody. The chromogen solution consisted of fast red dye and naphthol phosphate in Tris HCl buffer. A red insoluble product was deposited at sites of primary antibody binding. The tissue sections were rinsed, counterstained briefly in Gill hematoxylin, rinsed in bluing solution, and dehydrated through graded alcohols to xylene; a coverslip was applied with crystal mount, and tissue sections were examined microscopically. The immunohistochemical staining procedure was validated with positive and negative avian control tissues, including sections of blood vessels.
Evaluation of immunohistochemical staining—The degree of immunohistochemical staining was graded by intensity. A negative reaction had no discernible staining. For the purposes of this study, tissues had either positive or negative immunohistochemical staining for C psittaci antigen.
Severity of lesions—A full histologic description was given for each atherosclerotic lesion, and a lesion severity score from 1 to 4 was assigned to each study bird. Lesions with a score of 1 were considered mild and had isolated macrophages and intracellular lipid only. Lesions assigned a score of 2 were moderate and had macrophages with intracellular and extracellular lipid and a core of extracellular lipid. Marked lesions assigned a score of 3 had intracellular and extracellular lipids, fibrosis, calcification, and chondroid or osseus metaplasia. The most severe lesions with a score of 4 had all of the criteria for a score of 3 as well as surface defects such as hematomas, hemorrhage, and thrombus formation.
Statistical analysis—Exact logistic regression was used to evaluate the association of potential risk factors with atherosclerosis. Variables that were associated with atherosclerosis in the univariable analysis (P < 0.20) were considered eligible for inclusion in the multivariable analysis. Multivariable model selection began with a maximum model containing all eligible explanatory variables, and individual variables were removed in a manual stepwise process on the basis of the P value. Variables were retained in the multivariable model if they had a value of P < 0.10 or if their removal resulted in a change of > 10% to the other coefficients, indicating that their inclusion was necessary to achieve control of confounding. The most appropriate form of continuous predictor variables was determined by comparing Akaike information criterion for models that incorporated linear and categorized versions of the variables. The form of the predictor variable that yielded the lowest Akaike information criterion was preferred. Data were screened for outliers by examining plots of the predicted probabilities versus Δ χ2, Δ deviance, and Δ beta statistics on the basis of the maximum likelihood estimation of the final multivariable model. Goodness of fit was evaluated with the Hosmer-Leme-show goodness-of-fit test. Clinicopathologic values of study birds and control birds were compared by means of the Mann-Whitney test. All testing was performed assuming a 2-sided alternative hypothesis, and values of P < 0.05 were considered significant. Analyses were performed with commercially available statistical software.c
Results
Review of the medical records for the study period yielded information on 42 birds in which a diagnosis of atherosclerosis had been made; however, 11 birds were excluded from this study because medical records were not available or complete or appropriate vascular tissue was not available for immunohistochemical staining (Figures 1 and 2). The 31 birds included in the study ranged from 5 months to 32 years of age at the time of diagnosis (median age, 10.6 years). Sixteen were ≥ 10 years of age. Thirteen (42%) birds were male, 17 (55%) were female, and 1 (3%) was of unknown sex. Species represented were lovebirds (Agapornis sp; n = 2), budgerigars (Melopsittacus undulatus; 3), Amazon parrots (Amazona sp; 3), cockatoos (Cacatua sp; 2), grey-cheeked parakeets (Brotogeris Pyrrhopterus; 2), cockatiels (Nymphicus hollandicus; 5); quaker-monk parakeets (Myiopsitta monachus; 2), macaws (Ara sp; 3), African grey parrots (Psittacus erithacus; 4), and conures (Aratinga sp; 4); there was 1 unknown species termed parrot. The study population was further classified as 14 (45%) New World species, 6 (19%) African species, and 10 (32%) Australasian species on the basis of their natural origin, with 1 unknown species (3%). Most of the birds in the study were on seed-based pet bird diets.
Photomicrograph of a section of the aorta from a pet psittacine bird with atherosclerosis. Notice that the intima contains deposition of lipid and infiltrates of macrophages and small lymphocytes. Intralesional Chlamydophila psittaci antigen is identified by deposition of red chromogen. Avidin-biotin complex immunoperoxidase staining, primary mouse anti–C psittaci monoclonal antibody, fast red chromogen, and Gill hematoxylin counterstain; bar = 25 μm.
Citation: Journal of the American Veterinary Medical Association 240, 12; 10.2460/javma.240.12.1474
Photomicrograph of a section of the aorta and atheroma from a pet psittacine bird with atherosclerosis. Notice that the intima contains deposition of lipid and acicular clefts or cholesterol crystals. Intralesional C psittaci antigen is identified by deposition of red chromogen. Avidin-biotin complex immunoperoxidase staining, primary mouse anti–C psittaci monoclonal antibody, fast red chromogen, and Gill hematoxylin counterstain; bar = 25 μm.
Citation: Journal of the American Veterinary Medical Association 240, 12; 10.2460/javma.240.12.1474
Univariable risk factor analysis—The distribution of bird characteristics was summarized for study birds (n = 31) and control birds (31; Table 1). Positive immunohistochemical staining of blood vessels for C psittaci antigen and age of birds were both significantly associated with atherosclerosis. The odds of atherosclerosis was 8 times as high for birds with positive immunohistochemical staining for C psittaci antigen as for those with negative immunohistochemical staining. Compared with adult birds (2 to 10 years old), juvenile birds (0 to 1 years old) were less likely (OR, 0.17) to have atherosclerosis and geriatric birds (≥ 11 years old) were more likely (OR, 2.8) to have atherosclerosis. The odds of developing atherosclerosis were approximately 3 times as high for African and South American species, compared with the odds for Australasian species, although this was not a significant association. No other potential risk factors were significantly associated with atherosclerosis in the univariable analysis.
Univariate evaluation of risk factors for atherosclerosis in pet psittacine birds in a retrospective case-control study of 31 study birds and 31 control birds.
| Variable | No. of control birds* | No. of study birds* | OR (95% CI) | P value† |
|---|---|---|---|---|
| Chlamydophila psittaci antigen‡ | ||||
| Negative | 26 | 11 | Referent | < 0.001 |
| Positive | 5 | 18 | 8.2 (2.2–35.7) | |
| Species origin | ||||
| Australasian | 19 | 10 | Referent | 0.093 |
| South American | 8 | 14 | 3.2 (0.91–12.5) | |
| African | 4 | 6 | 2.8 (0.52–16.8) | |
| Age category (y) | ||||
| 2–10 | 16 | 14 | Referent | 0.016 |
| 0–1 | 17 | 1 | 0.17 (0.003–1.6) | |
| ≥ 11 | 6 | 15 | 2.8 (0.76–11.4) | |
| Sex | ||||
| Female | 14 | 17 | Referent | 0.622 |
| Male | 14 | 13 | 0.77 (0.24–2.4) | |
| Unknown | 3 | 1 | 0.28 (0.005–4.0) | |
| Body weight | ||||
| Ideal | 16 | 15 | Referent | 0.284 |
| Underweight | 11 | 7 | 0.68 (0.17–2.6) | |
| Overweight | 1 | 4 | 4.1 (0.35–223) | |
| Illness type | ||||
| Nonspecific | 12 | 6 | Referent | 0.134 |
| Gastrointestinal | 7 | 6 | 1.7 (0.31–9.4) | |
| Neurologic | 3 | 11 | 6.8 (1.2–53.4) | |
| Reproductive | 3 | 4 | 2.6 (0.32–23.7) | |
| Respiratory | 5 | 4 | 1.6 (0.22–10.9) | |
| Admission | ||||
| Appointment | 17 | 12 | Referent | 0.204 |
| Emergency | 13 | 18 | 2.0 (0.66–6.5) | |
| Diet | ||||
| Seed | 19 | 25 | Referent | 0.186 |
| Pellet | 3 | 1 | 0.26 (0.005–3.5) | |
| Seed and pellet | 5 | 1 | 0.16 (0.003–1.6) | |
| Other | 1 | 1 | 0.76 (0.009–62.8) | |
Number of study birds and control birds differ across comparisons because of missing information.
Exacttest of independence.
Negative or positive immunohistochemical staining of blood vessels for C psittaci antigen.
CI = Confidence interval.
Multivariable risk factor analysis—Results of the multivariable logistic regression analysis are summarized (Table 2). Only the result of immunohistochemical staining for C psittaci antigen was significantly associated with atherosclerosis in the final model. After adjusting for age, species origin, and type of illness, the odds of having atherosclerosis was 7 times as high for birds with positive immunohistochemical staining for C psittaci antigen, compared with that of birds with negative immunohistochemical staining. Illness type was grouped into 3 categories on the basis of results of the univariable analysis, and although this variable did not achieve significance at the 5% level (P = 0.072), birds with signs of neurologic disease were approximately 8 times as likely to have atherosclerosis as were birds with nonspecific signs of illness. Age and species origin were not significantly associated with atherosclerosis in the final multivariable model but were included because they were important confounders of immunohistochemical staining results and illness type. Age was included as a continuous variable because this yielded a stronger association with atherosclerosis than when it was included as a categorical variable. The continuous form of age also provided a lower value of Akaike information criterion when preliminary models were constructed with maximum likelihood estimation. The model provided a good fit to the data as measured by the Hosmer-Lemeshow goodness-of-fit test (χ26df = 5.20; P = 0.518). Only 1 bird, a 4-year-old cockatiel with atherosclerosis, respiratory illness, and negative immunohistochemical staining for C psittaci antigen, had both Δ χ2 and Δ deviance values that were > 4 in magnitude. This bird was retained in the model because it had plausible covariate values and did not exert a large influence on the model estimates.
Multivariable exact logistic regression model for the evaluation of risk factors for atherosclerosis in pet psittacine birds with complete covariate information (study birds, n = 27; control birds, 28).
| Variable | OR (exact 95% CI) | P value* |
|---|---|---|
| C psittaci antigen† | ||
| Negative | Referent | 0.012 |
| Positive | 7.0 (1.4–43.4) | |
| Species origin | ||
| Australasian | Referent | 0.544 |
| South American | 2.5 (0.38–20.3) | |
| African | 1.7 (0.15–24.6) | |
| Age (y) | 1.09 (0.97–1.24) | 0.178 |
| Illness type | ||
| Nonspecific | Referent | 0.072 |
| Gastrointestinal, reproductive, or respiratory | 1.8 (0.24–14.1) | |
| Neurologic | 8.4 (0.86–102) | |
Multivariable tests are based on score statistics.
Negative or positive immunohistochemical staining of blood vessels for C psittaci antigen.
See Table 1 for remainder of key.
Clinicopathologic findings—Clinicopathologic values were available for a subset of the study birds and control birds (Table 3). Study birds and control birds differed significantly only with respect to plasma cholesterol concentrations. Median plasma cholesterol concentration of study birds (421 mg/dL) was significantly (P = 0.008) higher than that of control birds (223 mg/dL).
Summary of clinicopathologic values in pet psittacine birds evaluated in a retrospective case-control study to identify risk factors for atherosclerosis.
| Variable | Control birds | Study birds | |||
|---|---|---|---|---|---|
| No. | Median (IQR) | No. | Median (IQR) | P value* | |
| Hematologic values | |||||
| Hct (%) | 23 | 45 (28–50) | 21 | 47 (45–50) | 0.161 |
| Total WBC count (× 103/μL) | 24 | 8.0 (4.1–10) | 21 | 9.0 (6.1–12) | 0.411 |
| Heterophil (%) | 24 | 74 (62–81) | 20 | 78 (69–82) | 0.389 |
| Lymphocyte (%) | 24 | 23 (17–28) | 20 | 22 (17–31) | 0.869 |
| Monocyte (%) | 24 | 1.5 (1.0–2.5) | 20 | 1.0 (1.0–2.0) | 0.807 |
| Plasma biochemical values | |||||
| Total protein (g/dL) | 20 | 2.6 (2.2–3.0) | 21 | 2.7 (2.5–3.6) | 0.256 |
| Aspartate aminotransferase (U/L) | 20 | 470 (226–761) | 20 | 408 (270–557) | 0.534 |
| Creatine kinase (U/L) | 20 | 536 (207–1,372) | 19 | 651 (325–2,057) | 0.238 |
| Glucose (mg/dL) | 19 | 302 (220–325) | 21 | 285 (244–345) | 0.766 |
| Calcium (mg/dL) | 20 | 8.5 (7.9–9.2) | 21 | 8.7 (7.8–10.2) | 0.375 |
| Phosphorus (mg/dL) | 20 | 4.5 (2.8–6.1) | 21 | 4.5 (3.3–7.4) | 0.764 |
| Uric acid (mg/dL) | 20 | 4.7 (3.3–7.7) | 22 | 4.9 (3.0–18.2) | 0.650 |
| Cholesterol (mg/dL) | 11 | 223 (144–250) | 11 | 421 (233–906) | 0.008 |
Mann-Whitney test.
IQR = Interquartile range.
Lesion severity scoring—Lesion severity scores were available for the study birds. Twenty-one of the 31 (68%) study birds had a score of 3, and 4 (13%) had a score of 4, indicating that 25 (81%) birds had marked to severe atherosclerotic lesions. Two of the 4 birds with the most severe lesions were New World species (macaws). Two of the 31 (6%) birds had a score of 1, and 4 (13%) had a score of 2.
Of the birds with marked and severe lesions (score, 3 and 4), 8 were male and 16 were female (the sex of 1 bird was unknown). All 8 Australasian species birds with a severity score of 3 were female. All 4 birds with severe lesions (score, 4) and 11 of 21 with marked lesions (score, 3) had positive immunohistochemical staining for C psittaci antigen. Two of the 4 birds with moderate lesions (score, 2) and 1 of the 2 birds with mild lesions (score, 1) had positive immunohistochemical staining for C psittaci antigen.
Discussion
In this study, 2 significant risk factors for developing atherosclerosis in pet psittacine birds were identified; they were positive immunohistochemical staining for C psittaci antigen in blood vessels and high plasma cholesterol concentrations. The presence of C psittaci antigen in atheromas was identified with a special immunohistochemical stain; 18 of 29 study birds had positive immunohistochemical staining for C psittaci antigen, whereas only 5 of 31 control birds had positive immunohistochemical staining for C psittaci antigen. Immunohistochemistry is used to identify proteins in cells of a tissue section via the principle of antibodies binding specifically to antigens in biological tissues.32 Immunohistochemistry is useful in the diagnosis of certain cancers and has had increasing diagnostic value in veterinary medicine. The reported prevalence of C pneumoniae in atherosclerotic tissue in human patients appears to depend on the detection system used (eg, immunohistochemistry or PCR assay). In the study reported here, immunostaining was used because of the availability of a novel stain specific for Chlamydia sp antigen. Immunostaining has been used in other studies6,33 to evaluate atheromas in avian tissues; however, the methods used in those studies differed from that of the present study.
Antemortem testing for C psittaci infection is available by several testing methods, including PCR assay and elementary body agglutination; however, each method has variable sensitivity and specificity for detection of infection. Also, the intracellular nature of organisms in the Chlamydiaceae family can preclude detection in some cases. Because suspicion of current C psittaci infection in the case population of the present study was low, birds did not undergo antemortem testing for infection. Information on whether any of the birds had undergone previous testing for C psittaci infection was unavailable. Based on clinical experience, positive immunohistochemical staining for C psittaci antigen is uncommon even in birds with high index of suspicion for infection. From the present study, it was not possible to determine whether previous or concurrent C psittaci infection at the time of necropsy was the cause of atherosclerosis; however, the identification of C psittaci antigen in atheromas indicates that infection may be a risk factor.
In the study reported here, the finding that elevation of plasma cholesterol concentration was a significant risk factor for atherosclerosis in pet psittacine birds was not surprising, given the known association in human patients. In the present study, total plasma cholesterol concentrations were evaluated. Investigation and validation of lipid profiles in psittacine birds to determine high-density lipoprotein and low-density lipoprotein concentrations would be beneficial if these tests were to become commercially available. In human patients, low high-density lipoprotein, high low-density lipoprotein, and high triglyceride concentrations are risk factors for atherosclerosis. Reference ranges were determined for each class of lipoprotein and triglycerides in Amazon parrots of 1 study.34
Although age is considered a common risk factor for many diseases, birds can have much longer life spans than other pets and vascular diseases can mimic those of human patients. Age was associated with atherosclerosis in the univariable analysis, but it was not a significant predictor in the multivariable model. Although not significant, the point estimate for the effect of age in the multivariable model suggested that the odds of atherosclerosis increased by approximately 9%/y (OR, 1.09).
Another factor evaluated in the study reported here were clinical signs of pet psittacine birds at the time of initial evaluation. Birds with signs of neurologic disease were an estimated 8 times as likely to have atherosclerosis as were birds with nonspecific signs of illness, although this finding was not significant. Vascular disease in birds can manifest as stroke-like behavior, weakness and falling, lameness, dull mentation, and sudden death.
Limited information was available on diet for the study population of the present study from the birds' clinical history obtained from the owners. Most birds were consuming a commercial seed or seed-based diet. Historically, these types of diets have been implicated in chronic nutritional and vitamin deficiencies and considered high in fat, which can lead to obesity. Body weight was not identified as an important risk factor for atherosclerosis; however, no avian standards exist (eg, body mass index for human patients), and body weights of birds were based on acceptable ranges for each species.35
Classification of atherosclerotic lesions by stage or grade is important because it helps distinguish clinically relevant lesions that may relate to the cause of death from lesions that may be incidental. In humans, the American Heart Association–recommended classification describes lesion types I to VIII with increasing severity, where types I to III are usually clinically silent.3 The use of this classification system has not been investigated in parrots, and other lesion classification schemes have been established in birds.6,33 In the population of birds of the present study, 25 birds had severe lesions and had clinically related, relevant disease supporting valid risk factor determination. Results of a similar study33 revealed that 4 (of which 3 were psittacine birds) of 103 birds (92 Psittaciformes in the study) were positive for Chlamydophila organisms (3 for C psittaci) as determined with a PCR assay on paraffin-embedded atherosclerotic tissue samples; however, all 4 birds had negative immunohistochemical staining for C psittaci antigen. Two of the 4 birds (only 1 was a psittacine) had severe atherosclerosis on the basis of the lesion severity scoring scale used in that study, indicating that birds with severe atherosclerosis may be more likely to have an C psittaci infection.
Because control selection is recognized as a potential source of bias in case-control studies, control bird selection in the present study was based on several factors: available study birds meeting inclusion criteria during the study period, region of origin of study birds, convenience sampling, and information on all psittacine birds seen during the study period. A potential limitation of the present study could also involve the accuracy and specificity of the immunohistochemical stain used. Studies36,37 in human patients have shown that some immunohistochemical stains will cross react with nonchlamydial plaque constituents. Because a primary mouse anti–C psittaci monoclonal antibodyb was used in the present study, staining was thought to be specific for chlamydial antigens. The monoclonal antibody does not have cross-reactivity with other bacterial species, and immunohistochemical staining is specific for the chlamydial antigen. In the present study, no nonspecific staining of tissues was observed in the great vessels and several tissue samples had negative immunohistochemical staining. In the present study, another diagnostic testing method was not used to confirm the immunohistochemical staining results. Unfortunately, there is no perfectly accurate test for the diagnosis of C psittaci infection currently available.
Most importantly, the identification of risk factors should lead to methods of prevention or to a decrease in the risk of developing disease. Management of atherosclerosis requires improved clinical diagnosis, consideration of species predilection, and feeding strategies.2 Veterinarians should consider genetic basis, methods of exercise options, and attempts to minimize stress. Medical management also requires addressing secondary conditions such as vascular stenosis, pulmonary disease, and heart failure. Treatment to reduce body weight and nutritional imbalances and increase dietary α-linolenic acid and frequent monitoring of the patient's plasma cholesterol (and triglyceride) concentrations are vitally important. The use of cholesterol-lowering statin drugs appears to be clinically useful in avian patients. Other treatment options in human patients, such as plasma delipidation (direct filtration of cholesterol from the blood with transfusion back into the patient) and newer medications (eg, statins), which inhibit cholesterol synthesis in humans, may show promise in the future management of avian patients.
DNA in situ Hybridization Laboratory, Department of Pathology, College of Veterinary Medicine, University of Georgia, Athens, Ga.
Chlamydia monoclonal antibody 10R-C124A, Fitzgerald Industries International Inc, Concord, Mass.
Stata, version 11.0, StataCorp LP, College Station, Tex.
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