Cell Identification - College of American Pathologists

Cell Identification

Case History This peripheral blood smear is from a 1-month-old male infant with a three-week history of cough and difficulty

breathing. Laboratory data include: Bordetella pertussis antigen smear = positive; Bordetella pertussis culture = positive; WBC = 37.4 x 109/L; HGB = 13.6 g/dL; HCT = 39.9%; MCV = 96 fL; RDW = 16.7; Platelet = 416 x 109/L; Absolute lymphocyte count = 29.5 x 109/L.

(PERIPHERAL BLOOD, WRIGHT-GIEMSA)

Identification

Participants

No.

%

Evaluation

Teardrop cell (dacrocyte)

999

95.3

Educational

VPBS-02

The image is of a teardrop shaped red blood cell (dacrocyte) and was correctly identified by 95.3% of participants. A teardrop shaped erythrocyte is typically normocytic in size but may be microcytic. Teardrop cells contain a single, short or long, blunted or rounded end. Unlike red blood cells with a short sharp pointed end which are artifacts of slide preparation, a true teardrop cell should taper into a blunt tip. Teardrop cells are typically associated with an abnormality of the spleen or bone marrow which causes the erythrocyte to stretch in order to continue circulating through the splenic cords or bone marrow sinuses.

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Case History

This peripheral blood smear is from a 1-month-old male infant with a three-week history of cough and difficulty breathing. Laboratory data include: Bordetella pertussis antigen smear = positive; Bordetella pertussis culture = positive; WBC = 37.4 x 109/L; HGB = 13.6 g/dL; HCT = 39.9%; MCV = 96 fL; RDW = 16.7; Platelet = 416 x 109/L; Absolute lymphocyte count = 29.5 x 109/L. (PERIPHERAL BLOOD, WRIGHT-GIEMSA)

Identification

Participants

No.

%

Evaluation

Lymphocyte

1038

99.0

Educational

VPBS-03

The image is of a lymphocyte and was correctly identified by 99.0% of participants. A lymphocyte may range in size from 7 to 15 ?m and typically has a high N:C ratio. Most lymphocytes are small with round to oval nuclei; however occasional normal lymphocytes may be medium in size due to increased cytoplasm. The nuclear chromatin is diffusely dense or coarse and clumped. Nucleoli, if present, are small and inconspicuous.

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Case History This peripheral blood smear is from a 1-month-old male infant with a three-week history of cough and difficulty

breathing. Laboratory data include: Bordetella pertussis antigen smear = positive; Bordetella pertussis culture = positive; WBC = 37.4 x 109/L; HGB = 13.6 g/dL; HCT = 39.9%; MCV = 96 fL; RDW = 16.7; Platelet = 416 x 109/L; Absolute lymphocyte count = 29.5 x 109/L.

(PERIPHERAL BLOOD, WRIGHT-GIEMSA)

Identification

Participants

No.

%

Evaluation

Basophil, any stage

1044

99.6

Educational

VPBS-04

The image is of a basophil that was correctly identified by 99.6% of the participants. Basophils are round to oval and are 10 to 15 ?m in size. Basophils characteristically contain a number of coarse, densely stained granules that vary in color from blue-black to red-purple. These granules are present throughout the cell and vary in size and shape and can obscure visualization of the nucleus.

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Case History This peripheral blood smear is from a 1-month-old male infant with a three-week history of cough and difficulty

breathing. Laboratory data include: Bordetella pertussis antigen smear = positive; Bordetella pertussis culture = positive; WBC = 37.4 x 109/L; HGB = 13.6 g/dL; HCT = 39.9%; MCV = 96 fL; RDW = 16.7; Platelet = 416 x 109/L; Absolute lymphocyte count = 29.5 x 109/L.

(PERIPHERAL BLOOD, WRIGHT-GIEMSA)

Identification

Participants

No.

%

Evaluation

Eosinophil, any stage

1047

99.9

Educational

VPBS-05

The image is of an eosinophil and was correctly identified by 99.9% of participants. Mature eosinophils are round to oval and are 10 to 15 ?m in size. Eosinophils contain characteristic coarse, orange-red granules of uniform size which typically do not overlie the cell nucleus. The nucleus is usually segmented into two to three lobes separated by a thin filament of chromatin. The chromatin is typically dense and compact. The nuclear lobes are equal in size and exhibit the same nuclear characteristics as neutrophils.

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Case History This peripheral blood smear is from a 1-month-old male infant with a three-week history of cough and difficulty

breathing. Laboratory data include: Bordetella pertussis antigen smear = positive; Bordetella pertussis culture = positive; WBC = 37.4 x 109/L; HGB = 13.6 g/dL; HCT = 39.9%; MCV = 96 fL; RDW = 16.7; Platelet = 416 x 109/L; Absolute lymphocyte count = 29.5 x 109/L.

(PERIPHERAL BLOOD, WRIGHT-GIEMSA)

Identification

Participants

No.

%

Evaluation

Lymphocyte, reactive (to include

566

54.1

plasmacytoid and immunoblastic

forms)

Lymphocyte

246

23.5

Monocyte

106

10.1

Lymphoma cell (malignant), includes

63

6.0

Hairy cell and Sezary cell

Educational

Educational Educational Educational

VPBS-06

The image is of a lymphocyte or reactive lymphocyte and was correctly identified by 77.6% of participants. A lymphocyte may range in size from 7 to 15 ?m with variable, typically high N:C ratio. The nucleus in the current cell shows an unusual deeply cleaved nucleus; however maintains the dense clumped chromatin. Normal lymphocytes may occasionally contain an indented nucleus which are more frequently seen in infants and children. This cell contains the characteristic findings of Bordetella pertussis lymphocytosis including the deeply clefted nucleus, coarse chromatin, small size and high to moderate N:C ratio.

Kathryn Rizzo, DO, PhD Hematology and Clinical Microscopy Resource Committee

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Discussion

Bordetella pertussis is a highly contagious bacteria which causes acute upper respiratory tract infection and the well-known epidemic disease, whooping cough (pertussis). Pertussis occurs worldwide, numbering over 48 million cases a year. Although vaccinations in the United States have decreased the incidence, pertussis infections can occur in previously vaccinated adolescents and adults and then can be transmitted to unvaccinated or partially immunized infants and children.

Infection is transmitted from person-to-person by airborne transmission and typical pertussis infection is divided into three symptomatic stages. The first stage is the catarrhal stage which may last for several weeks. During this stage the person may have symptoms similar to a mild cold. The second stage is the paroxysmal stage where the patient experiences the characteristic severe coughs and "whooping" of air with the rapid inspiration of breath into the lungs. During this time, the patient develops a lymphocytosis. The last stage is the convalescent stage. Pertussis infection may be detected via culture, direct fluorescent antibody testing, serologic testing, and molecular testing by PCR methods.

The lymphocytosis seen in pertussis infection usually occurs in infected infants and children and is typically in the range of 10.0 x 109/L but may reach levels of 30.0 x 109/L or higher. The lymphocytes are predominantly T cells with a normal CD4 to CD8 ratio. Additionally, there may be an increase in monocytes, neutrophils and other lymphocytes such as B cells and NK (natural killer) cells. The lymphocytosis and overall increased white blood cell count is thought to be due to a decrease in the cell membrane protein known as L-selectin. L-selectin allows for migration of leukocytes out of the blood and into peripheral tissue and is integral in lymphocyte migration to lymph nodes. The decreased amount of this protein thus causes an accumulation of lymphocytes in the blood stream.

Benign Causes of Lymphocytosis

Benign causes of an absolute lymphocytosis can be subclassified into two main categories based on the morphology of the lymphocytes. The first category includes benign lymphocytosis with activated/reactive morphology (i.e. pleomorphic lymphocytosis). These lymphocytes are activated lymphocytes with varying morphology and size. The lymphocytes are typically large and have more abundant cytoplasm with radiating basophilia often seen at the periphery of the cell cytoplasm. The chromatin is typically somewhat more dispersed than a small lymphocyte. The nucleus may be round, oval, or irregular, and inconspicuous to distinct nucleoli may be seen in large immunoblasts. The main cause for this type of lymphocytosis is viral infections, many of which are due to Epstein-Barr virus (EBV) infection. (Figure 1) However other viruses may cause an infectious mononucleosis-like syndrome including cytomegalovirus (CMV), adenovirus and acute human immunodeficiency virus (HIV) infection. Additional causes of an activated/reactive appearing lymphocytosis include drug reaction, post-vaccination, chronic inflammatory disorders (e.g. autoimmune disorders), and other infectious agents.

The second category includes benign lymphocytosis with a mature or non-activated/non-reactive morphology. This type of lymphocytosis is typically composed of normal sized lymphocytes with high nuclear to cytoplasmic (N/C) ratios and mature coarse chromatin. A common cause for this type of lymphocytosis is Bordetella pertussis infection in children, as seen in our current case. (Figure 2) In pertussis infection the mature appearing lymphocytes may contain the characteristic deeply cleaved nuclei.

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Figure 1. Activated lymphocyte in EBV infection

Figure 2. Bordetella pertussis lymphocyte

Additional benign causes for a non-activated lymphocytosis include transient stress lymphocytosis and less commonly, persistent polyclonal B cell lymphocytosis and acute infectious lymphocytosis. Transient (acute) stress lymphocytosis may be seen in patients with acute medical conditions including myocardial infarction (heart attacks) and trauma. Typically the lymphocytosis is mild to moderate and composed mostly of small mature lymphocytes with occasional large granular lymphocytes. Persistent polyclonal B cell lymphocytosis has distinctive clinical findings, typically occurring in asymptomatic young to middle aged women who are smokers. However there are rare reports seen in neonates and infants. These lymphocytes differ from a pertussis infection as they are predominantly of B cell lineage but share similar morphological features such as high N/C ratio, mature chromatin and nuclear clefting.

Neoplastic lymphocytosis with cleaved nuclei

Mature B cell lineage leukemias/lymphomas

A number of neoplastic disorders may cause lymphocytosis with morphological features similar to our current case. Chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) is a common cause for lymphocytosis in the adult population. The cells are typically small to slightly larger than a normal lymphocyte and contain mature, coarse chromatin. (Figure 3) Occasional cases may have neoplastic lymphoid cells with cleaved, indented nuclei. A characteristic finding in circulating follicular lymphoma cells is nuclear folding, irregularity and cleaving. (Figure 4) Follicular lymphoma cells are small to slightly larger than a normal lymphocyte and contain scant to moderate cytoplasm with moderately clumped chromatin and occasional small nucleolus. Mantle cell lymphoma is another B cell lineage lymphoma composed of lymphoma cells which are small to medium in size. (Figure 5) These lymphoma cells have moderately clumped chromatin and nuclei which may be folded or cleaved.

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Figure 3. Chronic Lymphocytic Leukemia

Figure 4. Follicular Lymphoma

Figure 5. Mantle cell Lymphoma

T cell lineage lymphomas T cell lymphomas are less common than B cell lymphomas; however they should also be considered in the differential diagnosis of a non-activated lymphocytosis with cleaved/irregular nuclei. The lymphoma cells in Sezary Syndrome are typically small with condensed chromatin and convoluted nuclei imparting a "cerebriform" appearance. A more uncommon lymphoma known as adult T cell leukemia/lymphoma contains lymphoma cells with markedly irregular, lobulated nuclear membranes.

Acute lymphoblastic leukemia An important differential to consider in our current pediatric case is acute B or T lymphoblastic leukemia. Lymphoblasts may be difficult to distinguish from normal lymphoid cells when they are small in size, contain mature appearing moderately granular chromatin, lack distinguishable nucleoli and have round to irregular nuclei. (Figure 6) However, even in these cases, occasional lymphoblasts with more typical morphology such as larger size, finer chromatin and distinct nucleoli will be seen. Thus, it is helpful to scan the slide on low power to evaluate the entire range of lymphoid morphology.

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