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Indications and Techniques for Blood Transfusion in Birds
Topics in Medicine and Surgery
Indications and Techniques for Blood Transfusion in Birds Filipe Martinho, DVM
Abstract There is growing medical care related to emergency presentations and procedures used to treat companion and wild birds. These critical care procedures (e.g., blood transfusion) can be life-saving. To maximize the beneficial effects of blood transfusions administered to avian patients, it is necessary to have an understanding of avian hematology and erythropoiesis, recognize clinical conditions in which one performs a blood transfusion, know the proper procedures and techniques, and rapidly identify possible adverse reactions. Copyright 2009 Elsevier Inc. All rights reserved. Key words: blood; cross matching; erythropoiesis; hemolysis; transfusion
M
ature avian erythrocytes or red blood cells (RBCs) are nucleated, usually with an oval form and a centrally placed nucleus containing uniformly clumped chromatin. With Giemsaand Romanowsky-based staining techniques, the nucleus appears deep purple and the cytoplasm light pink. Immature avian erythrocytes have a more rounded shape and nucleus, with more lightly clumped chromatin, and the cytoplasm stains light blue. The presence of some of these immature cells in the peripheral blood is normal and indicative of RBC regeneration, which is often described as polychromasia or polychromatophilia. A slight degree of RBC anisocytosis also may be considered a normal finding in healthy birds.1,2 Erythropoiesis takes place in the bone marrow and comprises 8 sequential stages of cell development with progressive morphological and staining changes: rubriblasts (or erythroblasts), prorubricytes, basophilic rubricytes, initial polychromatic rubricytes, late polychromatic rubricytes, rubricytes, polychromatic RBCs (or reticulocytes), and mature RBCs.1-4 Erythropoiesis is controlled by a number of different factors, including oxygen concentration in tissues and hormones. Hypoxia stimulates the production and release of erythropoietin, a glycoprotein produced in the kidneys, which has a direct positive
112
effect on erythropoiesis. Other hormones, such as androgens and adrenocorticoids, also contribute to the development of RBCs. In contrast, polycythemia tends to suppress erythropoiesis. Avian RBCs have a shorter half-life (25 to 45 days) than mammal cells, and, because of this more rapid turnover, birds usually show a higher degree of polychromasia than mammals.1-4 Blood volume in avian species is estimated between 4% and 8% of body weight (grams), with younger birds having a higher blood volume than adults. Compared with mammals, avian blood is more viscous because the RBCs are larger and less deformable. Blood density is mostly influenced by the concentration of plasma proteins, although birds
From the Faculdade de Medicina Veterinária, Universidade Lusófona de Humanidades e Tecnologia, Lisbon, Portugal. Address correspondence to: Filipe Martinho, DVM, Faculdade de Medicina Veterinária, Universidade Lusófona de Humanidades e Tecnologia, Lisbon, Portugal. E-mail: martinhfilipe@ gmail.com © 2009 Elsevier Inc. All rights reserved. 1557-5063/09/1802-$30.00 doi:10.1053/j.jepm.2009.04.001
Journal of Exotic Pet Medicine, Vol 18, No 2 (April), 2009: pp 112–116
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Blood Transfusion in Birds
have a lower blood albumin concentration and oncotic pressure than mammals.2,5 Evaluation of RBCs of a patient can be accomplished by measuring a hematocrit or packed cell volume (PCV), total RBC count, hemoglobin concentration, estimation of mean corpuscular volume and mean corpuscular hemoglobin, reticulocyte count, and RBC morphology. The PCV is calculated by centrifugation of microhematocrit capillary tubes and varies between 35% and 55% among different species of birds. A PCV less than 35% suggests anemia, whereas a PCV greater than 55% is indicative of dehydration or polycythemia. Within and between species, PCV can vary with age, gender, hormones, and other physiologic factors (e.g., males and older birds tend to have a higher PCV).1,3 The total RBC count in birds is usually estimated by manual methods, but recent advances in flow cytometric analyzer technology have allowed properly adjusted machines to be used. Total RBC counts in birds are generally lower (1.5 to 4.5 ⫻ 106 cells/ L) than those reported for mammals. RBC counts can be influenced by a number of different physiologic parameters, age, sex, hormones, hypoxia, and environmental factors.1,3 The mean corpuscular volume is estimated based on the following formula: PCV/RBC ⫻ 10; whereas the mean corpuscular hemoglobin can be determined by: hemoglobin concentration/RBC ⫻ 10. These indices are useful when classifying the type of anemic condition affecting the patient.1,2 RBC morphology is an important component of the erythron and can provide information regarding erythroid function. A slight polychromasia (between 0.4% and 6.78% in psittacines is considered normal) and anisocytosis are expected to be seen in avian blood smears. Other important changes that can occur and should be measured include poikilocytosis, nuclear abnormalities (e.g., fragmentation, binucleation, pyknosis, abnormal shapes, Howell-Jolly bodies), basophilic stippling of the cytoplasm, spherocytosis, Heinz body formation, and erythroplastids.1,3 Reticulocytes are immature cells and can be counted in blood smears stained with new methylene blue, which stains residual cytoplasmic RNA dark blue. Mature RBCs can also show some basophilic clumps surrounding the nucleus, so cell morphology should be used to characterize the reticulocytes. Reticulocyte counts generally vary between 1% and 5% in normal birds, with higher levels suggestive of RBC regeneration. Because the reticulocyte count gives a more precise estimate of RBC regeneration than polychromasia, it is the preferred method for characterizing regeneration.1-3
Anemia and Shock in Birds Anemia is classically defined by a decrease in PCV, RBC, and hemoglobin, and can be classified in 3 groups: blood loss anemia, regenerative anemia, and nonregenerative anemia.6 Blood loss anemia appears nonregenerative in the acute phase of blood loss but becomes progressively regenerative. Possible causes include trauma (e.g., lacerations, broken pin feathers, exposed fractures), parasites (e.g., ticks, Dermanyssus spp., coccidia), primary coagulopathies (rare in birds), coagulopathies secondary to toxicosis (e.g., rodenticide intoxication, aflatoxicosis) or nutrition (e.g., vitamin K deficiencies), and other nonspecific causes (e.g., gastrointestinal tract ulceration, ulcerated neoplasia, organ rupture). Regenerative or hemolytic anemia can be caused by hemoparasites (e.g., Plasmodium spp., Aegyptianella spp.), sepsis, toxicosis (e.g., mustard, oil intoxication), immune-mediated disease,7 and thermal injuries (e.g., secondary to burns). Nonregenerative anemia can develop quickly in birds because of their short RBC half-life and may be caused by chronic infections (e.g., tuberculosis, avian chlamydiosis, aspergillosis), neoplasia, hypothyroidism, hyperestrogenism, toxicosis (e.g., heavy metal intoxication, aflatoxicosis), nutritional imbalances (e.g., protein, iron, vitamin B deficiencies), leukemia (e.g., lymphoid leukemia, erythroblastosis), myeloblastosis, and hepatopathies or nephropathies. Birds appear more capable of coping with acute blood loss and accommodate better to chronic blood loss than mammals. It has been shown that chickens can return to a normal PCV 72 hours after removing 30% of their blood volume,8 whereas pigeons have been shown to return to a normal PCV 7 days after removing 60% of their blood volume without adverse clinical signs.9 Birds have this amazing capacity of physiologic accommodation because the extravascular space quickly replaces any lost vascular fluids, and the bone marrow has the ability to mobilize large numbers of immature RBCs. The absence of some autonomic responses to hypovolemic shock in birds also increases survival.8,10-16 Hypovolemic shock can develop either when there is a decreased blood volume or inadequate distribution of blood flow. This decreased blood flow can be absolute (e.g., hemorrhage, coagulopathy) or relative (e.g., dehydration, polycythemia). When a bird is in a considerable hypovolemic state (e.g., more than 30% loss of blood volume), there is also a decrease in the blood pressure and activation of baroreceptors and the vasomotor center in the medulla oblongata. This leads to activation of the sym-
114 pathetic nervous system, which can lead to vasoconstriction of peripheral veins and arterioles, increased heart rate and myocardial contraction, increased production and release of catecholamines, activation of the juxtaglomerular cells in the kidneys, release of renin, and activation of the renin-angiotensin-aldosterone system. The ultimate goal of all of these mechanisms is to restore normal blood pressure. Parallel to these mechanisms, mobilization of large numbers of immature RBCs from the bone marrow occurs. Shock can be divided in 3 distinct stages. In the initial/compensatory stage (e.g., loss of less than 20% of the blood volume), birds present with tachycardia, normal or increased blood pressure, strong and fast pulses, and a capillary refill time (CRT) of less than 1 second. The avian patient’s response to crystalloid fluid administration is usually uneventful. In the next stage, or early decompensatory phase (e.g., continuous loss of blood or a decrease of 25% to 30% of blood volume), birds present in a hypothermic state, exhibiting clinical signs such as cold extremities and skin, tachycardia, normal or decreased blood pressure, pale mucous membranes, increased CRT, and mental depression. To treat these patients, aggressive fluid therapy with both crystalloid and colloid products is warranted. The final stage, or late decompensatory phase, occurs when there is significant blood loss (more than 60% of blood volume), the autonomic neuroendocrine responses to shock become ineffective, and there is generalized organ failure. Common clinical signs associated with birds in the decompensatory phase are severe hypotension, pale or cyanotic mucous membranes, absent CRT, weak or absent pulses, hypothermia, oliguria or anuria, pulmonary edema, stupor or coma, and cardio-respiratory arrest. Usually avian patients that present in the decompensatory phase have a very poor prognosis, despite all therapeutic efforts.3,12
Indications and Special Considerations for Blood Transfusion Blood transfusions are rarely used in the early stages of hypovolemic shock, except in cases with severe and acute hemorrhage or some coagulopathies (e.g., rodenticide toxicosis). It is, however, indicated when there is a lack of RBCs, thrombocytes, coagulation factors, albumin, or antithrombine. Objectively, a blood transfusion should be provided when an avian patient is suffering from a loss of more than 20% of blood volume, when PCV is less than 20%, or in surgical patients with chronic ane-
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mia. For each case, it is important to assess the patient’s ability to cope with the stress of administering the blood transfusion and the potential adverse reactions to the blood transfusion. A blood transfusion should not be performed in normovolemic or dehydrated patients, in a bird with mild/moderate chronic anemia but otherwise healthy, in anemic and dehydrated patients (more than 7% dehydration), or in anemic patients with hypoproteinemia.10,11,14-16 In the domestic chicken, approximately 28 blood groups and 3 different blood group systems (B, L, and N) have been described.17 Blood groups have also been studied in a few galliforme and anseriforme species, but are unknown for the vast majority of birds currently treated in veterinary practices. As a safe measure, cross-matching should be performed
Table 1. Cross-Matching Procedure for Birds 1. Centrifuge (3500 rpm, 1 min) 1 drop of whole blood without anticoagulant to obtain serum and 1 drop of blood with EDTA to obtain RBCs from both the donor and recipient. 2. RBC washing: re-suspend the RBCs in 0.5 mL of saline solution, centrifuge 1 min, and discard supernatant. Repeat this procedure twice. 3. Re-suspend the washed RBCs in 0.5 mL of saline solution to obtain an RBC solution. 4. Major cross-matching: in a tube, place 2 drops of patient serum and 1 drop of donor RBC solution. 5. Minor cross-matching: in a tube, place 2 drops of donor serum and 1 drop of patient RBC solution. 6. Controls: one tube with 1 drop of patient serum and 2 drops of patient RBC solution and another tube with 1 drop of donor serum and 2 drops of donor RBC solution. 7. Incubate all tubes 15 min at 37°C. 8. Centrifuge all tubes for 15 sec. 9. Look for macroscopic agglutination and hemolysis. Re-suspend the RBCs and place 1 drop on a slide, apply a coverslip, and look for signs of microscopic agglutination under a microscope. Agglutination should be differentiated from rouleaux formation, which is a plasma-related phenomenon where RBCs are clumped in piles by effect of electrostatic forces. If a cross-match is compatible, the RBCs are individually distributed and there are no signs of hemolysis. Abbreviation: EDTA, ethylenediamine tetraacetic acid.
Blood Transfusion in Birds
before every blood transfusion, especially when a heterologous donor is used or when a bird is receiving a second transfusion (Table 1). Whenever possible, homologous (same species) transfusions are preferred, because the donor’s RBCs survive longer in a conspecific. Frequently, it is impossible to perform a homologous transfusion, so a heterologous (different species) transfusion must be performed. Because birds do not have preformed antibodies for blood groups, the first heterologous transfusion is usually safe, although hemolysis of donor RBCs always leads to some physiologic stress.13,14,16 The efficiency of homologous/heterologous transfusions has been studied in a small number of avian species. When measured, the mean donor RBC halflife was approximately 7 days; 1 day in homologous transfusions in pigeons but only 12 hours in heterologous pigeon-red tailed hawk (Buteo jamaicensis) transfusions18; 10 to 16.8 days in homologous transfusions in cockatiels but only 0.1 to 2.6 days in heterologous transfusions6; and 8.5 days in homologous transfusions in Aratinga conures and 4.5 days in heterologous transfusions in 2 different species of Aratinga.19 Based on this evidence, heterologous transfusions should be ideally performed between related genera to increase the likelihood of RBC survival.
Collection and Administration of Blood Products Blood should be collected from a healthy donor bird (about 1% body weight and replaced with crystalloid fluids), preferably with the bird under general inhalant anesthesia, and mixed with appropriate anticoagulants. Citrate-phosphate-dextrose-adenosine or sodium citrate anticoagulants are preferred because they are rapidly metabolized by the recipient and are less likely to cause coagulation problems. The anticoagulant product should be mixed with blood in a 1:9 ratio. If these preferred anticoagulant products are not available, heparin can be used by adding 0.25 mL of heparin to 10 mL of blood. Fresh blood is preferred for the transfusion, because prolonged storage leads to an increased release of potassium and there is generally no adequate method for storing avian blood.14,20 The avian recipient should receive 10 to 20 mL of blood/kilogram, warmed to body temperature, to keep the patient’s PCV above 25% and blood pressure above 90 mm Hg. Blood can be administered intravenously or intraosseously with a blood filter by slow bolus injection (5 to 10 minutes) or by an injection pump over a 4-hour period. Blood transfusions can be performed as a first-line treatment in
115 patients with severe hemorrhage, but are usually given after stabilization of the avian patient with oxygen, crystalloid, and/or colloid fluid therapy and other supportive care.10-12,13-15 As an alternative to blood or blood products, Oxyglobin (Biopure Corp., Cambridge, MA USA) can be used. This is a hemoglobin-based oxygen carrier, comprised of purified bovine hemoglobin in a modified lactated Ringer’s solution. It acts as a colloid, delivering oxygen to tissues and having a vasoconstrictor effect, and helps to counteract the physiologic consequences of shock. Oxyglobin is not immunogenic; therefore, no cross-matching is required and a blood filter is not needed for administration. The Oxyglobin product can be stored at room temperature for 3 years, but after the bag is compromised it should be used within a 24-hour period to prevent the potential of meta-hemoglobin formation. Birds should receive 5 mL/kg of Oxyglobin intravenously or intraosseously by bolus injection over some minutes.10,12,13,15,16 Despite all these advantages, Oxyglobin does have potential side effects. Because of the red color of the product, it can stain the mucous membranes red and alter the color of the plasma, which can be confused with hemolysis. Recently, some concerns have been raised in dogs and humans regarding the effectiveness of Oxyglobin’s ability to deliver oxygen to tissues because of its vasoconstrictive effect and the reduced cardiac output that accompanies shock. There have also been deaths reported after Oxyglobin administration in dogs that developed a subsequent autoimmune hemolytic anemia and gastrointestinal irritation. Humans have been diagnosed with hypertension after Oxyglobin treatment. Although no adverse reactions in birds have been reported, caution should be used when Oxyglobin is administered.21 Crystalloid fluid therapy with or without iron-dextran administration can also be an effective treatment in a hypovolemic patient if blood or oxyglobin are not available. It has been demonstrated that, in pigeons and quail, lactated Ringer’s solution or saline solution administration in conjunction with iron-dextran therapy was as effective as a blood transfusion to restore a normal PCV.22,23 When conducting a blood transfusion in an avian patient, it is important to monitor the animal for potential side effects. The most common side effects associated with blood transfusions in vertebrates include hemolysis of donor RBCs, fever, urticaria, and anaphylaxis. These conditions are often difficult to assess in birds, but have been reported in patients receiving multiple blood transfusions. Fortunately,
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many of these problems can be prevented by crossmatching before a transfusion. Regurgitation has also been reported in birds secondary to hypervolemia (e.g., blood is given too quickly or in excess) and can be prevented by administering the blood transfusion over a 4-hour period.13,15
10. 11. 12.
Conclusion 13.
Although blood transfusions are seldom performed in birds, they can be life-saving in certain situations. Proper evaluation of the avian patient and selection of individuals that meet the criteria for blood administration is important, as well as recognition of advantages and limitations of blood transfusion and alternative therapies.
References 1. 2.
3. 4. 5. 6. 7. 8. 9.
Campbell TW (ed): Avian Hematology and Cytology (ed 2). Ames, IA, Iowa State University Press, 1995 Campbell TW: Hematology, in Ritchie BW, Harrison GJ, Harrison LR (eds): Avian Medicine: Principles and Application. Lake Worth, FL, Wingers Publishing, pp 176-198, 1994 Mitchell EB, Johns J: Avian hematology and associated disorders. Vet Clin North Am (Exotic Anim Pract) 11:501-522, 2008 Campbell TW, Ellis KC (eds): Avian and Exotic Hematology and Cytology (ed 3). Ames, IA, Blackwell, 2007 Sturkie PD, Griminger P: Body fluids: blood, in Sturkie PD (ed): Avian Physiology. New York, NY, SpringerVerlag, pp 102-114, 1986 Pendl H: Anemia in birds—an overview. Proceedings of the Annual Conference of the European Association of Avian Veterinarians, 160-162, 2001 Johnston MS, Son TT, Rosenthal KL: Immune-mediated hemolytic anemia in an eclectus parrot. J Am Vet Med Assoc 230:1028-1031, 2007 Ploucha JM, Scott JB, Ringer RK: Vascular and hematologic effects of hemorrhage in the chicken. Am J Physiol 240:H9-H17, 1981 Finnegan VM, Daniel GB, Ramsey EC: Evaluation of
14. 15. 16. 17. 18.
19.
20.
21.
22.
23.
whole blood transfusions in domestic pigeons (Columba livia). J Avian Med Surg 11:7-14, 1997 Bowles H, Lichtenberger M, Lennox A: Emergency and critical care of pet birds. Vet Clin North Am (Exotic Anim Pract) 10:345-394, 2007 Campbell T: Stabilization of the emergency avian patient. Exotic Pet Pract 2:89-90, 1997 Lichtenberger M: Shock and cardiopulmonary-cerebral resuscitation in small mammals and birds. Vet Clin North Am (Exotic Anim Pract) 10:275-291, 2007 Matos R, Morrisey JK: Emergency and critical care of small psittacines and passerines. Semin Avian Exot Pet Med 14:90-105, 2005 Morrisey JK: Avian transfusion medicine. Exotic Pet Pract 4:65-66, 1999 Speer BL: Stabilizing companion birds in an emergency. Vet Med December:832-842, 2005 Lichtenberger M: Transfusion medicine in exotic pets. Clin Tech Small Anim Pract 19:88-95, 2004 Dietert MF, Taylor RL, Dietert RR: Avian blood groups. Poult Sci Rev 4:87-105, 1992 Sandmeier P, Stauber EH, Wardrop KJ, et al: Survival of pigeon red blood cells after transfusion into selected raptors. J Am Vet Med Assoc 204:427-429, 1994 Degernes LA, Harrison LD, Smith DW: Autologous, homologous and heterologous red blood cell transfusions in conures of the genus Aratinga. J Avian Med Sur 13:10-14, 1999 Morrisey JK, Hohenhaus AE, Rosenthal KL, et al: Comparison of three media for the storage of avian whole blood. Proceedings of the Association of Avian Veterinarians, 279-290, 1997 Meyer R: Current topics in fluid therapy: Oxyglobin, in Gleed RD, Ludders JW (eds): Recent Advances in Veterinary Anesthesia and Analgesia: Companion Animals. Ithaca, NY, International Veterinary Information Service (IVIS), 2008. Available from: URL:http:// www.ivis.org/advances/Anesthesia_Gleed/meyer/ chapter_frm.asp?LA⫽1 Schindler SL, Gildersleeve RP, Thaston JP, et al: Hematological response of haemorrhaged Japanese quail after blood volume replacement with saline. Comp Biochem Physiol 87A:933-945, 1987 Bos JH, Todd B, Tell LA: Treatment of anemic birds with iron dextran therapy, homologous and heterologous blood transfusions. Proceedings of the Annual Conference of the Association of Avian Veterinarians, 221-225, 1990
Indications and Techniques for Blood Transfusion in Birds Filipe Martinho, DVM
Abstract There is growing medical care related to emergency presentations and procedures used to treat companion and wild birds. These critical care procedures (e.g., blood transfusion) can be life-saving. To maximize the beneficial effects of blood transfusions administered to avian patients, it is necessary to have an understanding of avian hematology and erythropoiesis, recognize clinical conditions in which one performs a blood transfusion, know the proper procedures and techniques, and rapidly identify possible adverse reactions. Copyright 2009 Elsevier Inc. All rights reserved. Key words: blood; cross matching; erythropoiesis; hemolysis; transfusion
M
ature avian erythrocytes or red blood cells (RBCs) are nucleated, usually with an oval form and a centrally placed nucleus containing uniformly clumped chromatin. With Giemsaand Romanowsky-based staining techniques, the nucleus appears deep purple and the cytoplasm light pink. Immature avian erythrocytes have a more rounded shape and nucleus, with more lightly clumped chromatin, and the cytoplasm stains light blue. The presence of some of these immature cells in the peripheral blood is normal and indicative of RBC regeneration, which is often described as polychromasia or polychromatophilia. A slight degree of RBC anisocytosis also may be considered a normal finding in healthy birds.1,2 Erythropoiesis takes place in the bone marrow and comprises 8 sequential stages of cell development with progressive morphological and staining changes: rubriblasts (or erythroblasts), prorubricytes, basophilic rubricytes, initial polychromatic rubricytes, late polychromatic rubricytes, rubricytes, polychromatic RBCs (or reticulocytes), and mature RBCs.1-4 Erythropoiesis is controlled by a number of different factors, including oxygen concentration in tissues and hormones. Hypoxia stimulates the production and release of erythropoietin, a glycoprotein produced in the kidneys, which has a direct positive
112
effect on erythropoiesis. Other hormones, such as androgens and adrenocorticoids, also contribute to the development of RBCs. In contrast, polycythemia tends to suppress erythropoiesis. Avian RBCs have a shorter half-life (25 to 45 days) than mammal cells, and, because of this more rapid turnover, birds usually show a higher degree of polychromasia than mammals.1-4 Blood volume in avian species is estimated between 4% and 8% of body weight (grams), with younger birds having a higher blood volume than adults. Compared with mammals, avian blood is more viscous because the RBCs are larger and less deformable. Blood density is mostly influenced by the concentration of plasma proteins, although birds
From the Faculdade de Medicina Veterinária, Universidade Lusófona de Humanidades e Tecnologia, Lisbon, Portugal. Address correspondence to: Filipe Martinho, DVM, Faculdade de Medicina Veterinária, Universidade Lusófona de Humanidades e Tecnologia, Lisbon, Portugal. E-mail: martinhfilipe@ gmail.com © 2009 Elsevier Inc. All rights reserved. 1557-5063/09/1802-$30.00 doi:10.1053/j.jepm.2009.04.001
Journal of Exotic Pet Medicine, Vol 18, No 2 (April), 2009: pp 112–116
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Blood Transfusion in Birds
have a lower blood albumin concentration and oncotic pressure than mammals.2,5 Evaluation of RBCs of a patient can be accomplished by measuring a hematocrit or packed cell volume (PCV), total RBC count, hemoglobin concentration, estimation of mean corpuscular volume and mean corpuscular hemoglobin, reticulocyte count, and RBC morphology. The PCV is calculated by centrifugation of microhematocrit capillary tubes and varies between 35% and 55% among different species of birds. A PCV less than 35% suggests anemia, whereas a PCV greater than 55% is indicative of dehydration or polycythemia. Within and between species, PCV can vary with age, gender, hormones, and other physiologic factors (e.g., males and older birds tend to have a higher PCV).1,3 The total RBC count in birds is usually estimated by manual methods, but recent advances in flow cytometric analyzer technology have allowed properly adjusted machines to be used. Total RBC counts in birds are generally lower (1.5 to 4.5 ⫻ 106 cells/ L) than those reported for mammals. RBC counts can be influenced by a number of different physiologic parameters, age, sex, hormones, hypoxia, and environmental factors.1,3 The mean corpuscular volume is estimated based on the following formula: PCV/RBC ⫻ 10; whereas the mean corpuscular hemoglobin can be determined by: hemoglobin concentration/RBC ⫻ 10. These indices are useful when classifying the type of anemic condition affecting the patient.1,2 RBC morphology is an important component of the erythron and can provide information regarding erythroid function. A slight polychromasia (between 0.4% and 6.78% in psittacines is considered normal) and anisocytosis are expected to be seen in avian blood smears. Other important changes that can occur and should be measured include poikilocytosis, nuclear abnormalities (e.g., fragmentation, binucleation, pyknosis, abnormal shapes, Howell-Jolly bodies), basophilic stippling of the cytoplasm, spherocytosis, Heinz body formation, and erythroplastids.1,3 Reticulocytes are immature cells and can be counted in blood smears stained with new methylene blue, which stains residual cytoplasmic RNA dark blue. Mature RBCs can also show some basophilic clumps surrounding the nucleus, so cell morphology should be used to characterize the reticulocytes. Reticulocyte counts generally vary between 1% and 5% in normal birds, with higher levels suggestive of RBC regeneration. Because the reticulocyte count gives a more precise estimate of RBC regeneration than polychromasia, it is the preferred method for characterizing regeneration.1-3
Anemia and Shock in Birds Anemia is classically defined by a decrease in PCV, RBC, and hemoglobin, and can be classified in 3 groups: blood loss anemia, regenerative anemia, and nonregenerative anemia.6 Blood loss anemia appears nonregenerative in the acute phase of blood loss but becomes progressively regenerative. Possible causes include trauma (e.g., lacerations, broken pin feathers, exposed fractures), parasites (e.g., ticks, Dermanyssus spp., coccidia), primary coagulopathies (rare in birds), coagulopathies secondary to toxicosis (e.g., rodenticide intoxication, aflatoxicosis) or nutrition (e.g., vitamin K deficiencies), and other nonspecific causes (e.g., gastrointestinal tract ulceration, ulcerated neoplasia, organ rupture). Regenerative or hemolytic anemia can be caused by hemoparasites (e.g., Plasmodium spp., Aegyptianella spp.), sepsis, toxicosis (e.g., mustard, oil intoxication), immune-mediated disease,7 and thermal injuries (e.g., secondary to burns). Nonregenerative anemia can develop quickly in birds because of their short RBC half-life and may be caused by chronic infections (e.g., tuberculosis, avian chlamydiosis, aspergillosis), neoplasia, hypothyroidism, hyperestrogenism, toxicosis (e.g., heavy metal intoxication, aflatoxicosis), nutritional imbalances (e.g., protein, iron, vitamin B deficiencies), leukemia (e.g., lymphoid leukemia, erythroblastosis), myeloblastosis, and hepatopathies or nephropathies. Birds appear more capable of coping with acute blood loss and accommodate better to chronic blood loss than mammals. It has been shown that chickens can return to a normal PCV 72 hours after removing 30% of their blood volume,8 whereas pigeons have been shown to return to a normal PCV 7 days after removing 60% of their blood volume without adverse clinical signs.9 Birds have this amazing capacity of physiologic accommodation because the extravascular space quickly replaces any lost vascular fluids, and the bone marrow has the ability to mobilize large numbers of immature RBCs. The absence of some autonomic responses to hypovolemic shock in birds also increases survival.8,10-16 Hypovolemic shock can develop either when there is a decreased blood volume or inadequate distribution of blood flow. This decreased blood flow can be absolute (e.g., hemorrhage, coagulopathy) or relative (e.g., dehydration, polycythemia). When a bird is in a considerable hypovolemic state (e.g., more than 30% loss of blood volume), there is also a decrease in the blood pressure and activation of baroreceptors and the vasomotor center in the medulla oblongata. This leads to activation of the sym-
114 pathetic nervous system, which can lead to vasoconstriction of peripheral veins and arterioles, increased heart rate and myocardial contraction, increased production and release of catecholamines, activation of the juxtaglomerular cells in the kidneys, release of renin, and activation of the renin-angiotensin-aldosterone system. The ultimate goal of all of these mechanisms is to restore normal blood pressure. Parallel to these mechanisms, mobilization of large numbers of immature RBCs from the bone marrow occurs. Shock can be divided in 3 distinct stages. In the initial/compensatory stage (e.g., loss of less than 20% of the blood volume), birds present with tachycardia, normal or increased blood pressure, strong and fast pulses, and a capillary refill time (CRT) of less than 1 second. The avian patient’s response to crystalloid fluid administration is usually uneventful. In the next stage, or early decompensatory phase (e.g., continuous loss of blood or a decrease of 25% to 30% of blood volume), birds present in a hypothermic state, exhibiting clinical signs such as cold extremities and skin, tachycardia, normal or decreased blood pressure, pale mucous membranes, increased CRT, and mental depression. To treat these patients, aggressive fluid therapy with both crystalloid and colloid products is warranted. The final stage, or late decompensatory phase, occurs when there is significant blood loss (more than 60% of blood volume), the autonomic neuroendocrine responses to shock become ineffective, and there is generalized organ failure. Common clinical signs associated with birds in the decompensatory phase are severe hypotension, pale or cyanotic mucous membranes, absent CRT, weak or absent pulses, hypothermia, oliguria or anuria, pulmonary edema, stupor or coma, and cardio-respiratory arrest. Usually avian patients that present in the decompensatory phase have a very poor prognosis, despite all therapeutic efforts.3,12
Indications and Special Considerations for Blood Transfusion Blood transfusions are rarely used in the early stages of hypovolemic shock, except in cases with severe and acute hemorrhage or some coagulopathies (e.g., rodenticide toxicosis). It is, however, indicated when there is a lack of RBCs, thrombocytes, coagulation factors, albumin, or antithrombine. Objectively, a blood transfusion should be provided when an avian patient is suffering from a loss of more than 20% of blood volume, when PCV is less than 20%, or in surgical patients with chronic ane-
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mia. For each case, it is important to assess the patient’s ability to cope with the stress of administering the blood transfusion and the potential adverse reactions to the blood transfusion. A blood transfusion should not be performed in normovolemic or dehydrated patients, in a bird with mild/moderate chronic anemia but otherwise healthy, in anemic and dehydrated patients (more than 7% dehydration), or in anemic patients with hypoproteinemia.10,11,14-16 In the domestic chicken, approximately 28 blood groups and 3 different blood group systems (B, L, and N) have been described.17 Blood groups have also been studied in a few galliforme and anseriforme species, but are unknown for the vast majority of birds currently treated in veterinary practices. As a safe measure, cross-matching should be performed
Table 1. Cross-Matching Procedure for Birds 1. Centrifuge (3500 rpm, 1 min) 1 drop of whole blood without anticoagulant to obtain serum and 1 drop of blood with EDTA to obtain RBCs from both the donor and recipient. 2. RBC washing: re-suspend the RBCs in 0.5 mL of saline solution, centrifuge 1 min, and discard supernatant. Repeat this procedure twice. 3. Re-suspend the washed RBCs in 0.5 mL of saline solution to obtain an RBC solution. 4. Major cross-matching: in a tube, place 2 drops of patient serum and 1 drop of donor RBC solution. 5. Minor cross-matching: in a tube, place 2 drops of donor serum and 1 drop of patient RBC solution. 6. Controls: one tube with 1 drop of patient serum and 2 drops of patient RBC solution and another tube with 1 drop of donor serum and 2 drops of donor RBC solution. 7. Incubate all tubes 15 min at 37°C. 8. Centrifuge all tubes for 15 sec. 9. Look for macroscopic agglutination and hemolysis. Re-suspend the RBCs and place 1 drop on a slide, apply a coverslip, and look for signs of microscopic agglutination under a microscope. Agglutination should be differentiated from rouleaux formation, which is a plasma-related phenomenon where RBCs are clumped in piles by effect of electrostatic forces. If a cross-match is compatible, the RBCs are individually distributed and there are no signs of hemolysis. Abbreviation: EDTA, ethylenediamine tetraacetic acid.
Blood Transfusion in Birds
before every blood transfusion, especially when a heterologous donor is used or when a bird is receiving a second transfusion (Table 1). Whenever possible, homologous (same species) transfusions are preferred, because the donor’s RBCs survive longer in a conspecific. Frequently, it is impossible to perform a homologous transfusion, so a heterologous (different species) transfusion must be performed. Because birds do not have preformed antibodies for blood groups, the first heterologous transfusion is usually safe, although hemolysis of donor RBCs always leads to some physiologic stress.13,14,16 The efficiency of homologous/heterologous transfusions has been studied in a small number of avian species. When measured, the mean donor RBC halflife was approximately 7 days; 1 day in homologous transfusions in pigeons but only 12 hours in heterologous pigeon-red tailed hawk (Buteo jamaicensis) transfusions18; 10 to 16.8 days in homologous transfusions in cockatiels but only 0.1 to 2.6 days in heterologous transfusions6; and 8.5 days in homologous transfusions in Aratinga conures and 4.5 days in heterologous transfusions in 2 different species of Aratinga.19 Based on this evidence, heterologous transfusions should be ideally performed between related genera to increase the likelihood of RBC survival.
Collection and Administration of Blood Products Blood should be collected from a healthy donor bird (about 1% body weight and replaced with crystalloid fluids), preferably with the bird under general inhalant anesthesia, and mixed with appropriate anticoagulants. Citrate-phosphate-dextrose-adenosine or sodium citrate anticoagulants are preferred because they are rapidly metabolized by the recipient and are less likely to cause coagulation problems. The anticoagulant product should be mixed with blood in a 1:9 ratio. If these preferred anticoagulant products are not available, heparin can be used by adding 0.25 mL of heparin to 10 mL of blood. Fresh blood is preferred for the transfusion, because prolonged storage leads to an increased release of potassium and there is generally no adequate method for storing avian blood.14,20 The avian recipient should receive 10 to 20 mL of blood/kilogram, warmed to body temperature, to keep the patient’s PCV above 25% and blood pressure above 90 mm Hg. Blood can be administered intravenously or intraosseously with a blood filter by slow bolus injection (5 to 10 minutes) or by an injection pump over a 4-hour period. Blood transfusions can be performed as a first-line treatment in
115 patients with severe hemorrhage, but are usually given after stabilization of the avian patient with oxygen, crystalloid, and/or colloid fluid therapy and other supportive care.10-12,13-15 As an alternative to blood or blood products, Oxyglobin (Biopure Corp., Cambridge, MA USA) can be used. This is a hemoglobin-based oxygen carrier, comprised of purified bovine hemoglobin in a modified lactated Ringer’s solution. It acts as a colloid, delivering oxygen to tissues and having a vasoconstrictor effect, and helps to counteract the physiologic consequences of shock. Oxyglobin is not immunogenic; therefore, no cross-matching is required and a blood filter is not needed for administration. The Oxyglobin product can be stored at room temperature for 3 years, but after the bag is compromised it should be used within a 24-hour period to prevent the potential of meta-hemoglobin formation. Birds should receive 5 mL/kg of Oxyglobin intravenously or intraosseously by bolus injection over some minutes.10,12,13,15,16 Despite all these advantages, Oxyglobin does have potential side effects. Because of the red color of the product, it can stain the mucous membranes red and alter the color of the plasma, which can be confused with hemolysis. Recently, some concerns have been raised in dogs and humans regarding the effectiveness of Oxyglobin’s ability to deliver oxygen to tissues because of its vasoconstrictive effect and the reduced cardiac output that accompanies shock. There have also been deaths reported after Oxyglobin administration in dogs that developed a subsequent autoimmune hemolytic anemia and gastrointestinal irritation. Humans have been diagnosed with hypertension after Oxyglobin treatment. Although no adverse reactions in birds have been reported, caution should be used when Oxyglobin is administered.21 Crystalloid fluid therapy with or without iron-dextran administration can also be an effective treatment in a hypovolemic patient if blood or oxyglobin are not available. It has been demonstrated that, in pigeons and quail, lactated Ringer’s solution or saline solution administration in conjunction with iron-dextran therapy was as effective as a blood transfusion to restore a normal PCV.22,23 When conducting a blood transfusion in an avian patient, it is important to monitor the animal for potential side effects. The most common side effects associated with blood transfusions in vertebrates include hemolysis of donor RBCs, fever, urticaria, and anaphylaxis. These conditions are often difficult to assess in birds, but have been reported in patients receiving multiple blood transfusions. Fortunately,
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many of these problems can be prevented by crossmatching before a transfusion. Regurgitation has also been reported in birds secondary to hypervolemia (e.g., blood is given too quickly or in excess) and can be prevented by administering the blood transfusion over a 4-hour period.13,15
10. 11. 12.
Conclusion 13.
Although blood transfusions are seldom performed in birds, they can be life-saving in certain situations. Proper evaluation of the avian patient and selection of individuals that meet the criteria for blood administration is important, as well as recognition of advantages and limitations of blood transfusion and alternative therapies.
References 1. 2.
3. 4. 5. 6. 7. 8. 9.
Campbell TW (ed): Avian Hematology and Cytology (ed 2). Ames, IA, Iowa State University Press, 1995 Campbell TW: Hematology, in Ritchie BW, Harrison GJ, Harrison LR (eds): Avian Medicine: Principles and Application. Lake Worth, FL, Wingers Publishing, pp 176-198, 1994 Mitchell EB, Johns J: Avian hematology and associated disorders. Vet Clin North Am (Exotic Anim Pract) 11:501-522, 2008 Campbell TW, Ellis KC (eds): Avian and Exotic Hematology and Cytology (ed 3). Ames, IA, Blackwell, 2007 Sturkie PD, Griminger P: Body fluids: blood, in Sturkie PD (ed): Avian Physiology. New York, NY, SpringerVerlag, pp 102-114, 1986 Pendl H: Anemia in birds—an overview. Proceedings of the Annual Conference of the European Association of Avian Veterinarians, 160-162, 2001 Johnston MS, Son TT, Rosenthal KL: Immune-mediated hemolytic anemia in an eclectus parrot. J Am Vet Med Assoc 230:1028-1031, 2007 Ploucha JM, Scott JB, Ringer RK: Vascular and hematologic effects of hemorrhage in the chicken. Am J Physiol 240:H9-H17, 1981 Finnegan VM, Daniel GB, Ramsey EC: Evaluation of
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whole blood transfusions in domestic pigeons (Columba livia). J Avian Med Surg 11:7-14, 1997 Bowles H, Lichtenberger M, Lennox A: Emergency and critical care of pet birds. Vet Clin North Am (Exotic Anim Pract) 10:345-394, 2007 Campbell T: Stabilization of the emergency avian patient. Exotic Pet Pract 2:89-90, 1997 Lichtenberger M: Shock and cardiopulmonary-cerebral resuscitation in small mammals and birds. Vet Clin North Am (Exotic Anim Pract) 10:275-291, 2007 Matos R, Morrisey JK: Emergency and critical care of small psittacines and passerines. Semin Avian Exot Pet Med 14:90-105, 2005 Morrisey JK: Avian transfusion medicine. Exotic Pet Pract 4:65-66, 1999 Speer BL: Stabilizing companion birds in an emergency. Vet Med December:832-842, 2005 Lichtenberger M: Transfusion medicine in exotic pets. Clin Tech Small Anim Pract 19:88-95, 2004 Dietert MF, Taylor RL, Dietert RR: Avian blood groups. Poult Sci Rev 4:87-105, 1992 Sandmeier P, Stauber EH, Wardrop KJ, et al: Survival of pigeon red blood cells after transfusion into selected raptors. J Am Vet Med Assoc 204:427-429, 1994 Degernes LA, Harrison LD, Smith DW: Autologous, homologous and heterologous red blood cell transfusions in conures of the genus Aratinga. J Avian Med Sur 13:10-14, 1999 Morrisey JK, Hohenhaus AE, Rosenthal KL, et al: Comparison of three media for the storage of avian whole blood. Proceedings of the Association of Avian Veterinarians, 279-290, 1997 Meyer R: Current topics in fluid therapy: Oxyglobin, in Gleed RD, Ludders JW (eds): Recent Advances in Veterinary Anesthesia and Analgesia: Companion Animals. Ithaca, NY, International Veterinary Information Service (IVIS), 2008. Available from: URL:http:// www.ivis.org/advances/Anesthesia_Gleed/meyer/ chapter_frm.asp?LA⫽1 Schindler SL, Gildersleeve RP, Thaston JP, et al: Hematological response of haemorrhaged Japanese quail after blood volume replacement with saline. Comp Biochem Physiol 87A:933-945, 1987 Bos JH, Todd B, Tell LA: Treatment of anemic birds with iron dextran therapy, homologous and heterologous blood transfusions. Proceedings of the Annual Conference of the Association of Avian Veterinarians, 221-225, 1990