Introduction

Interpretation of the avian and reptile hemogram can be complex and confusing. Extrinsic factors (season, temperature, habitat, diet, disease, stress, venipuncture site) and intrinsic factors (species, gender, age, physiologic status) may affect the hemogram numbers, distribution of the leukocytes, and the patient’s response to disease. In reptiles, low environmental temperatures may suppress and inhibit the immune response of the reptile, and lymphocyte counts in general are lower during the winter months and higher during the summer. Detailed reviews of abnormal changes seen in the hemogram in response to selected disease processes are presented for microscopic evaluation and interpretation.

Erythrocytes

Hypochromasia: Greater than 2+ may be a result of nutritional deficiencies leading to iron deficiency, lead toxicity, or chronic inflammatory disease.

Anisocytosis: Greater than 1+ with an increase of polychromasia suggests a regenerative anemia.

Poikilocytosis: Greater than 1+ is associated with erythrocytic regeneration. Anisocytosis and poikilocytosis may also be found in the post-hibernation reptile or those with severe inflammatory disease.

Basophilic stippling suggests a regenerative response but may also occur in reptiles with iron deficiency or less commonly, lead toxicity.

Perinuclear rings or clear irregular refractile spaces in the cytoplasm are artifacts in the erythrocyte because of the blood smear drying too slowly. Do not confuse these with gametocytes of Hemoproteus sp. or Plasmodium sp. Round to irregular basophilic inclusions (clumping endoplasmic reticulum) are frequently seen in the erythrocyte cytoplasm and are considered artifacts of slide preparation.

Anemia

The normal PCV in the avian patient ranges from 35–55% and 15–55% in the reptile patient. Values above this range with an elevated total protein suggest dehydration or erythrocytosis (polycythemia) with a normal or low total protein. Values below this range suggest anemia if hemodilution with lymph is not a factor. Anemia is classified according to pathophysiology and is termed hemolytic, hemorrhagic, or hypoplastic (depression anemia). Polychromasia in the reptiles is usually not graded as it is in the avian species, since polychromasia is less than 1% of the total erythrocyte population. In birds, polychromasia is graded 1–4+ to indicate a response, with 1+ considered normal in the non-anemic bird. The presence of increased reticulocytes and immature erythrocytes is evidence of active erythropoiesis. In reptiles, the erythrocyte regenerative response is slower than that of birds (1 week) and may take up to 4 months to return to normal due to the long lifespan of the erythrocyte and the long development from the rubriblast stage to the mature erythrocyte stage. In contrast to the avian patient, erythrocytes showing binucleation, anisokaryosis, or mitotic activity may suggest a regenerative process. However, severe inflammatory disease, malnutrition, and starvation must also be ruled out in the reptile patient. Immature reptiles or reptiles undergoing ecdysis may show an increase in polychromasia and immature erythrocytes.

Hemorrhagic anemia represents acute blood loss as seen with traumatic injuries. Birds regulate blood loss more efficiently than mammals. Diving and flying birds are more resistant to blood loss. Birds can rapidly shift extracellular fluid to the vascular space and have the capacity to mobilize large numbers of immature erythrocytes. Diving and migratory birds are more resistant to blood loss. In both avian and reptile patients, blood-sucking parasites and conditions such as coagulopathies or ulcerative lesions should be considered.

Hemolytic anemia is often due to systemic or hematogenous bacterial infections (sepsis), toxins (lead, zinc, aflatoxins, petroleum products, and certain plants), hemoparasites (Plasmodium and Aegyptianella), or infectious diseases. Lead or zinc may cause a mild to moderate anemia with evidence of regeneration. Hemolytic anemia with Heinz body anemia is associated with oil pollution and petroleum exposure. Immune-mediated disease associated with hemolysis and agglutination of the erythrocytes is rare and is primary or secondary to neoplasia, drug reactions, or infectious agents. This type of anemia usually presents with a strong response to the anemia with small round erythrocytes (spherocytes) and biliverdinuria due to extravascular hemolysis.

Hypoplastic anemia is a nonregenerative, normocytic, normochromic anemia. Often termed depression anemia, this is the most common anemia seen in the avian and reptile species. There is a decrease of erythropoiesis due to chronic inflammation or infection, drug reaction, renal or hepatic disease, and/or neoplasia.

Myeloid leukemia is characterized by abnormal erythrocytes, mitotic figures, and a large number of rubriblasts.

White Blood Cell Changes with Disease

Heterophils become active and toxic with systemic illness (usually bacterial etiology). The severity of the systemic illness affects the degree of toxic changes present and the number of immature heterophils released into the peripheral blood. The toxic changes seen in the heterophil include an increase in cytoplasmic basophilia, degranulation, abnormal granulation (rounding), and vacuolization. The immature heterophils consist of bands, metamyelocytes, myelocytes, and progranulocytes. A regenerative left shift is when the mature heterophils outnumber the immature heterophils, whereas a degenerative left shift reflects the predominance of immature heterophils. The latter can be a grave situation.

Lymphocytes show reactivity by developing a basophilic cytoplasm or forming scalloped edges. Inclusion body disease (IBD) viral inclusions can be seen in the snake lymphocyte and confirmed with H&E stain.

Plasma cells distended with clear to light blue round structures are termed Mott cells. The round structures are termed Russell bodies, which contain immunoglobulins.

Monocytes/azurophils become larger with exaggerated foamy cytoplasm. Cytoplasmic vacuoles and blebbing may occur with reactivity. A monocytosis suggests chronic or granulomatous inflammation. Monocytes have phagocytic capabilities and often engulf leukocytes and erythrocytes in response to anemia and infectious disease. They also actively engulf bacteria in a sepsis condition.

Melanomacrophages are considered a type of macrophage. This cell can be found in the peripheral blood of reptiles with severe inflammatory disease and may carry a guarded prognosis. Changes suggest nonspecific severe heterophilic inflammation with increased demand for macrophages, possible tissue necrosis, and systemic antigenic stimulation.

Eosinophils may increase with parasitic infections and stimulation of the immune system, while basophils may increase with parasitic and viral infections, wounds, and respiratory disease.

Thrombocytes play an active role in the response to disease. The presence of an increased number of immature thrombocytes is a regenerative response to excessive utilization. A decrease in thrombocytes suggests decreased bone marrow production or excessive peripheral utilization. This may be due to septicemia or DIC. Activated thrombocytes will appear as aggregated clusters of cells.

Leukocytosis

Stress leukograms show an absolute leukocytosis with moderate mature heterophilia and lymphopenia. The H/L ratio is a reliable indicator of stress. One must also identify the lymphocytic species when interpreting the leukogram. The cell morphology is considered normal.

Inflammatory leukograms are variable depending on the severity of the illness or condition. Heterophils should predominate, and monocytes/azurophils may be present. Significant increases in heterophils are associated with inflammatory diseases, stress, and neoplasia. Localized infections often show a mild inflammatory leukogram. Reptiles can stimulate a marked leukemoid response with high numbers of reactive monocytes/azurophils to an inflammatory disease. This can be confused with an emerging neoplastic disorder but will resolve itself with treatment of the underlying disease process.

Severe inflammatory disease in birds or reptiles who develop a marked leukocytosis with the presence of immature heterophils (bands, metaheterophils, myeloheterophils, progranulocytes) and toxicity suggests a bacterial infection or sepsis.

Lymphocytosis may occur with wound healing, parasitic infections, viral infections, and inflammatory disease. A lymphocytosis may also be present during the reptile ecdysis. The presence of reactive lymphocytes and plasma cells reflects stimulation of the immune system. Chronic lymphocytic leukemia is also a possibility with very elevated white blood counts (adult lymphocytes predominating and normal in appearance).

When atypical large lymphocytes with scalloped edges are present, an underlying neoplasia (lymphosarcoma) process needs to be considered. However, some viral infections that are considered inflammatory (West Nile virus) may also present with atypical lymphocytes. Blast leukemia reveals unclassified cells of the mononuclear line with immature cells predominating.

Leukopenia

Leukopenia reflects the consumption or decreased production of the peripheral leukocytes. The presence of an absolute heteropenia in conjunction with immature heterophils and toxicity represents overwhelming demand for heterophils in the periphery due to bacterial sepsis or viral disease. This is a degenerative response and carries a grave prognosis. Leukopenia with heteropenia, anemia, and thrombocytopenia suggest injury to the bone marrow. Lymphopenia may occur secondary to diseases that cause immunosuppression, chronic malnutrition, and chronic stress.

Table 1. Avian and reptile hematology guide

References

1.  Campbell T. Peripheral blood of birds. Peripheral blood of reptiles. Exotic Animal Hematology and Cytology. Oxford UK: Blackwell; 2015.

2.  Giordano N, Stefania L, Mattia B. Clinical hematology in reptilian species. Vet Clin Exot Anim. 2013;16:1–30.

3.  Nicole S, Rick A, Katherine AS. Diagnostic hematology of reptiles. J Clin Lab Med. 2011;31:87–108.

4.  Raffaella C, Lorenzo C. Overview of psittacine blood analysis and comparative retrospective study of clinical diagnosis, hematology and blood chemistry in selected psittacine species. Vet Clin Exot Anim. 2013;16:71–120.