1

Anemie

Uw hematologie analyzer en een anemische patiënt

Prof Dr. Erik Teske, Dip ECVIM-CADept Clin Scie Comp Animals

Utrecht University

Definition Anaemia

• Shortage of circulating red blood cells, or

decrease in normal quantity of hemoglobin

Clinical Effects:

- Inadequate tissue oxygenation

• pale mucous membranes

• weakness, inappetence, anorexia

• syncope

- Compensatory mechanisms

• tachypnea (particularly if forced to exercise)

• tachycardia (short and strong pulse)

Signs which may be associated with cause of

anaemia

• icterus

• bleeding (petechiae, ecchymoses, melena,

hematuria, hematomas)

• fever

• splenomegaly

Additional Clinical Symptoms:

Anaemia

Non-Regenerative Regenerative

Primary SecondaryBone Marrow Disease

Anaemia

Non-Regenerative Regenerative

Primary SecondaryBone Marrow Disease

Blood loss Hemolysis

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How to differentiate?

• Non-regenerative anaemia often chronic;

patient is reasonable well with low haematocrit

• Regenerative anaemia could be associated with

typical clinical signs:

• Icterus

• Bleedings

• Dark red urine, blood in faeces

• Tachycardia

• Dyspnoea

Further differentiation regenerative

vs non-regenerative anaemia

• Reticulocytes (percentage, absolute numbers)

• MCV, MCH, MCHC, RDW

• Blood smear: polychromasia, anisocytosis,

nucleated erythrocytes

Anisocytosis and Polychromasia Nucleated Erythrocytes

Nucleated erythrocytes (normoblasts), polychromasia,

anisocytosis

Reticulocytes

May Grünwald Giemsa (or

Hemacolor/DiffQuick):anisocytosis,

polychromasia, normoblast

Briljant Cresyl Blue (or

New Methylene Blue):reticulocytes

Red Blood Cell Parameters

• MCV: Mean Corpuscular Volume (= average size of RBC)

• MCH: Mean Corpuscular Haemoglobin (= average amount

of oxygen-carrying hemoglobin inside a RBC)

• MCHC: Mean Corpuscular Hemoglobin Concentration (=

average concentration of hemoglobin inside a RBC)

• RDW: Red cell Distribution Width (= variation in size of

RBCs)

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Anaemia

Non-Regenerative Regenerative

Primary SecondaryBone Marrow Disease

Blood loss Hemolysis

Non-Regenerative anaemia:

Primary Bone Marrow Disorders

• Myelophthisis (neoplasia)

• Myelofibrosis (secondary to other diseases)

• Myelosclerosis/osteosclerosis

• Myelodysplastic diseases

• Aplastic anemia (infectious: FeLV, Parvo,

Ehrlichia; Drug-related: estrogen, cytostatics)

• Pure Red Cell Aplasia (immune-mediated)

• Hemophagocytic syndrome

Non-regenerative anaemia:

Secondary Bone Marrow Disorders

• Chronic inflammatory disease (AID)

• Some endocrine diseases (Hypothyroidism,

Hypoadrenocorticism)

• Chronic renal failure with decreased

erythropoietin levels

• Deficiencies: Folic acid/Vit B12, Fe, Copper

• Low Reticulocyte count

• Absence of polychromasia and anisocytosis

• Normocytic-normochromic anaemia:

- MCV and RDW, MCH and MCHC: within ref range

• Microcytic-hypochromic anaemia:

- Fe deficiency

• Macrocytic-normochromic anaemia:

- Folic acid/Vit B12 deficiency

Non-regenerative anaemia

laboratory findings

Non-regenerative anemialaboratory findings

• Primary (bone marrow disorders): • Diagnosis by bone marrow evaluation + specific tests

• Leukopenia and/or thrombocytopenia may also occur

• Secondary: Laboratory findings of the primary disease• e.g. chronic renal failure: ↑ urea and creatinine;

• endocrinologic: abnormal endocrine tests

In addition:

Caveat:

• It takes a few days (2-4) to get a visible

response in peripheral blood after acute

bleeding or hemolysis

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Anaemia

Non-Regenerative Regenerative

Primary SecondaryBone Marrow Disease

Blood loss Hemolysis

Blood loss

• History

• Physical examination (petechia, bleeding, hemothorax/-abdomen)

• Bloodwork:

• Total protein/albumin

• Thrombocytes

• Clotting times (PT, APTT), fibrinogen

• Chronic blood loss => Iron deficiency => non-regenerative microcytic, hypochromic anemia, low ret% and high platelets

Hemolysis

• History (often acute, red urine, yellow feces,

geographic area, medication/intoxication)

• Physical examination (pale, icterus, tachycardia)

• Bloodwork:

• Ht, Reticulocytes, blood smear

• Serology, PCR

• Auto-agglutination

• Osmotic fragility

• Coombs test

Hemolytic Anaemia

1) Intravascular hemolysis• Rare: Oxidative damage (onions), Congenital

(phosphofructokinase deficiency)

• Infections

2) Extravascular hemolysis• Immune mediated, infections

3) Combination

Intravascular hemolysis:

- Parasites/infectious causes

- Microangiopathic/Vascular Endothelial Lesions

(Schistocytes)

- Oxidant damage (Heinz bodies)

- Congenital (e.g. pyruvate kinase (PK)/

phosphofructokinase (PFK) deficiencies)

- Neonatal iso-erythrolysis (cats)

- Other (e.g hypophosphataemia)

Extravascular haemolysis

- Physiological: (aged erythrocytes) removed by the

macrophage-monocyte system in the spleen

- Pathological: (Auto)antibodies are produced against

“normal” erythrocytes that are phagocytosed by the

spleen =>

IMMUNE-MEDIATED HEMOLYTIC ANAEMIA

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Erythrophagocytosis in spleen

Immune-mediated Hemolytic

Anaemia

- Primary/Idiopathic (unknown mechanisms)

- Secondary to:

• Infectious agents (bacterial, viral, parasitic)

• Neoplasia

• Drugs/insecticides/vaccines/neonatal isoerythrolysis

WILL CAUSE THE APPEARANCE OF

ABNORMAL ANTIGENS ON THE ERYTHROCYTE

CELL MEMBRANE

Methods of hematology analysis

• Manual cell count

• Hematology analyzers

• Microscopic examination

Manual cell count Types of hematology analyzers

• Microcapillary/Buffycoat analysis

• Impedance/coulter principle

• Laser flowcytometry

6

Microcapillary/Buffycoat

analysisBuffycoat analysis

(QBC/VetAutoRead)

Buffycoat analysis

• Ht, Hb, and WBC: good correlation with professional analyzers

• Thrombocytes cat and horse: poorer correlation

• Reticulocytes: bad correlation

• Leukopenia often not recognized

• Eosinophils: often combined with lymphocytes/monocytes

Impedance/coulter principle(Vet ABC, Medonic)

Sample

Dilution

Lysis Isotonic

WBC Hgb RBC MCV Plt MPV

Hct

MCHC

(calculated)

Laser flowcytometry

• Laser light is scattered by cell

• Scattering dependent on different

characteristics

• Detectors measure absorbance under different

angles

Laser flowcytometrybed-side instruments (e.g. IDEXX LaserCyte)

7

Haematologic Measurements

Results in a few

seconds: 5000 cells!

Analist 10 minutes;

100 cells…

BUT: Analyzers

cannot replace good

laboratory person!!

High aangle detector

(5o - 15o)

Low angle detector

(2o-3o)

670nm

Laser

Diode

High aangle detector

(5o - 15o)

Low angle detector

(2o-3o)

670nm

Laser

Diode

High aangle detector

(5o - 15o)

Low angle detector

(2o-3o)

670nm

Laser

Diode

670nm

Laser

Diode

Question:

Do we still need microscopic blood

examination since we have modern

hematology analyzers?

Advantage blood smear evaluation

• Better judgment of morphology of cells

• Control cell count and differentiation

• Quality check

• Banded granulocytes cannot be recognized

on analyzers

• Nucleated erythrocytes

• Mast cells

• Parasites

• Leukemic cells

Case 1: Golden Retriever, male, 9 years old

• 3 days of fever

• depression

• Anorexia

• first time you are seeing this dog

Physical examination

• Some pale mucous membranes

• A few petechiae in the mouth

• CRT is less than 1 sec

• Pulse is 90/min and strong

• Temp 38.5 C

Problem list

• Pale membranes

• Petechiae

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Differential diagnosis Pale membranes

• Shock: not very likely (slow and strong pulse, CRT <1)

• Anaemia:

- Blood loss: No history of external blood loss

Check thorax and abdomen [ ]

- Haemolysis

- Non-regenerative

Plan: CBC, Ret%

Differential diagnosis Petechiae

• Thrombocytopenia

– Increased consumption

– Decreased production

– Destruction

• Thrombocytopathy

• (Vessel wall abnormalities)

• (Von Willebrand’s disease)Plan: Platelets numbers

Diagnostic approach:

Initial step:

Ht, Ret%, leukocytes+differentiation, platelets

Results LasercyteR:

Ht: 0.18 l/l, Ret 3.0%, Ret Abs 63.4 x109/l, Leukocytes count 2.3x109/l, Platelets 25.000

Microscopic evaluation blood smear: low neutro’s and no platelet clumps

RETICULOCYTE PRODUCTION INDEX: CALCULATIONS

- Corrected reticulocyte percentage (CRP)

CRP= % reticulocytes x PCV of sample/normal PCV

- Reticulocyte Production Index (RPI)

RPI= CRP/Maturation Index(MI) PCV MI (days)0.45 1

(values for dogs) 0.35 1.5

0.25 20.15 2.5

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- Corrected reticulocytes percentage (CRP)

CRP= 15% x 0.15/0.45 = 5%

- Reticulocyte Production Index (RPI)

RPI= 5/2.5 =2

Dog with PCV=0.15L/L and reticulocytes =15%

RETICULOCYTE PRODUCTION INDEX: PRACTICAL EXAMPLE

INTERPRETATION

RPI > 3 Very good regeneration

RPI = 1-3 Good regeneration

RPI < 1 Inadequate regeneration

SPECIES VARIATIONS: IN CLINICALLY HEALTHY ANIMALS

-Dogs. Low number of reticulocytes (<1%), aggregate only

- Cats. Two types of reticulocytes:

• aggregate: blue stained coarse clumping(0.5% of erythrocytes)

• punctate: small, blue stained dots (1-10%).

- Ruminants and horses. Virtually no reticulocytesin blood.

Feline Reticulocytes

PunctateAggregate

maturation

NRBC Mature

24 hours < 10 daysRBC

Feline Reticulocytes

PunctateAggregate

maturation

NRBC Mature

24 hours < 10 daysRBC

SPECIES VARIATION: in anaemicanimals

- Canine. Strong reticulocyte response in regenerative

anaemias.

Aggregated reticulocytes (indicate recent response)

- Feline.

Punctated reticulocytes (indicate response to anaemia

occurring 1-2 weeks previously)

- Ruminants and horses. Reticulocytes may not appear

even in very severe anaemias

Results:

• Ht: 0.18 l/l, Ret 3.0%, Leukocytes count 2.3x109/l, Platelets 25.000

• CRP= 3% x 0.18/0.45 = 1.2

• RPI = 1.2/MI (2) = 0.6

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(differential) diagnosis

• Non-regenerative anaemia

• Leukopenia

• Thrombocytopenia

• => Pancytopenia

Differential diagnosisPancytopenia

• - myeloproliferative and lympho-proliferative disorders

• - ehrlichiosis

• - toxic bone marrow depression (estrogens/cytostatics)

• - myelofibrosis

• - osteosclerosis

Diagnostic approach:

• - additional history

• - palpation testis

• - bone marrow aspiration

• Enlarged R testis => FNA: Sertoli cell tumor

• bone marrow aspiration: aplastic bone marrow.

• Reticulocytosis was defined as an absolute reticulocyte count

(Ret#) above the upper reference interval (reticulocytes > 110.0 x 109 /l in dogs and >50.0 x 109/l in cats).

• The prevalence of RWA is 4.4% in dogs (492/11087) and 3.1% in cats (124/3956).

• 1.5% of the dogs (7/458) and 1.8% of the cats (2/111) with

RWA were apparently healthy, the rest were diseased

• Mortality rate was 29.7% in dogs (136/458) and 37.8% in cats

(42/111)

Case #2

Cocker spaniel, male, 3 years old

Lethargic since 1 week, sometimes vomiting.

Pale mucous membranes, Pulse 150/min

Bloodwork In-House analyzer

• Ht 0.114 [0.42-0.61]

• RBC 1.31 [6.2-8.9]

• MCV 82.0 [63.5-72.9]

• MCH 1.63 [1.37-1.57]

• MCHC 22.7 [20.5-22.4]

• Thromb 19 [144-603]

• Leukocytes 24.4 [4.5-14.6]

• Lymphocytes 5.6 [0.8-4.7]

• Neutrophils 16.2 [3.0-11.0]

• Eosinophils [0.0-1.6]

• Basophils [0.0-0.1]

• Monocytes 2.6 [0.0-0.9]

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Bloodwork In-House analyzer

• Ht 0.114 [0.42-0.61]

• RBC 1.31 [6.2-8.9]

• MCV 82.0 [63.5-72.9]

• MCH 1.63 [1.37-1.57]

• MCHC 22.7 [20.5-22.4]

• Thromb 19 [144-603]

• Leukocytes 24.4 [4.5-14.6]

• Lymphocytes 5.6 [0.8-4.7]

• Neutrophils 16.2 [3.0-11.0]

• Eosinophils [0.0-1.6]

• Basophils [0.0-0.1]

• Monocytes 2.6 [0.0-0.9]

No microscopic evaluation blood smear performed.

Problem list based on analyzer results

• Severe anemia

• Severe thrombocytopenia

• Neutrophilia

• Lymphocytosis

• Monocytosis

What if microscopic evaluation blood smear would have been performed?

Spherocytes (IHA)

macrophage

12

COMPARISON OF NORMAL LYMPHOCYTES AND NUCLEATED RED

BLOOD CELLS (NRBC)

lymphocyte - NRBC

What about the thrombocytopenia?

False low platelet numbers due to

aggregates

1) Leukocytosis

2) Neutrophilia, regenerative left shift

3) Monocytosis

4) No lymphocytosis

Conclusions Leukogram:

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1) Anisocytosis

2) Moderate to marked polychromasia

3) Nucleated erythrocytes

4) Marked spherocytosis

5) Occasional ghost cells

1.Aggregates: most likely normal platelet numbers

2.Some large platelets: young platelets

Conclusions erytron

Conclusions thrombon

Interpretation:

1) Inflammatory leukogram with tissue

necrosis and mild systemic toxic changes

2) Anemia is highly regenerative and spherocytic. => suspect Immune-

mediated hemolytic anemia (IHA).

Tests in IMHA

• Blood smear: presence of spherocytes

• Autoagglutination

• Osmotic fragility (long/short)

• Coombs test (direct/indirect)

Direct or with in-saline slide agglutination test

Auto-Agglutination

Macroscopic Microscopic

Whole blood in a hypotonic solution (0.55% NaCl)

Normal RBCs absorb water from the

hypotonic solution for osmotic

equilibrium and are distended but not

haemolyzed

Membranes of fragile RBCs(spherocytes, and those with enzyme

deficiencies or damaged by some drugs)

cannot withstand distension and are

haemolyzed

Erythrocyte Fragility Test: PrincipleImmune mediated hemolytic anemia

Osmotic fragility of erytrocytes

0.131.7

0.9285.3

0.263.4

0.395.1

0.4126.8

0.5158.5

0.6190.2

0.7221.9

0.8253.6

% NaClmOsm/l

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COOMBS TESTDetects antibodies directed at the erythrocyte membrane

False Positive:

• some chronic infections

• parasites (heartworms, haemobartonella)

• drugs (trimethoprim-sulfa)

• neoplasms

False Negative:

• in some cases of inadequate antibody production

The test is species-specific

Immune mediated hemolytic anaemia

anti-IgG-FcAnti-IgG-κλanti-IgM

Ery’s patientCoombs test

Determines presence erythrocyte-bound antibodies

-isotype-titer-temperature activity

(4 C / 37 C)

Immune mediated hemolytic anaemia

• A polyvalent Coombsserum

• B anti-dog IgG-κλ

• C anti-dog IgM

• D anti-dog IgG-Fc

• E control, 0.9% NaCl

• At 40C and 370C

Case #3

Labrador, male, 8 years old

Losing weight, some vomiting since a few weeks, some anorexia.

Pale/pink mucous membranes, pulse 100/min

Bloodwork (UVDL)

• Ht 0.25 [0.42-0.61]

• MCV 61.1 [63.5-72.9]

• MCH 1.31 [1.37-1.57]

• MCHC 19.1 [20.5-22.4]

• Ret% 0.5

• CHr 1.13 [1.43-1.71]

• Thromb 890 [144-603]

• Leukocytes 18.6 [4.5-14.6]

• Lymphocytes 0.7 [0.8-4.7]

• Neutrophils 16.0 [3.0-11.0]

• Bands 1.0 [0.0-0.3]

• Eosinophils 0.1 [0.0-1.6]

• Basophils [0.0-0.1]

• Monocytes 0.8 [0.0-0.9]

Bloodwork (UVDL)

• Ht 0.25 [0.42-0.61]

• MCV 61.1 [63.5-72.9]

• MCH 1.31 [1.37-1.57]

• MCHC 19.1 [20.5-22.4]

• Ret% 0.5

• CHr 1.13 [1.43-1.71]

• Thromb 890 [144-603]

• Leukocytes 18.6 [4.5-14.6]

• Lymphocytes 0.7 [0.8-4.7]

• Neutrophils 16.0 [3.0-11.0]

• Bands 1.0 [0.0-0.3]

• Eosinophils 0.1 [0.0-1.6]

• Basophils [0.0-0.1]

• Monocytes 0.8 [0.0-0.9]

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Findings:

1) Neutrophilia with left shift

(regenerated)

2) Lymphopenia

3) Hypo/microcytic anaemia

4) Thrombocytosis

5) Schistocytes

Interpretation:

1) Mild inflammation with

superimposed stress

2) Suspect Iron deficiency anemia

3) Reactive thrombocytosis

4) Mechanic trauma erythrocytes?

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Interpretation:

1) Mild inflammation with

superimposed stress

2) Suspect Iron deficiency anemia

3) Reactive thrombocytosis

4) Mechanic trauma erythrocytes?

Disadvantages classic parameters for diagnosing Fe deficiency

• Insensitive parameters

• Only abnormal in late Fe deficiency stage

• Respond to inflammatory diseases

• Require additional blood sampling

• Time consuming

Alternative: CHr

ADVIA®2120 Hematology System

Reticulocyte Analysis

Absorbance

RNA Content

High angle detector

(5o - 15o)

(Hb concentration)

Low angle detector

(2o-3o) (Volume)

670nm

Laser

Diode

Oxazine 750

RNA

Stain

Flow

Reticulocyte Parameters

High Angle (5-15 degrees)

Low

An

gle

(2

-3 d

egre

es)

Fe deficiency and response on Fe therapy

Red: Hb in erythrocytes

Blue: Hb in reticulocytes

Normal Hb content

17

CHr/Fe to diagnose Fe def

Parameter Sensitivity Specificity

CHr < 1.15 80% 92,1%

Serum Fe < 10,4 95,8% 92,3%

Fe def anemia due to

• Iron deficient diet? Rarely

• Chronic blood loss!

Leiomyosarcoma

Further diagnostic test: ultrasound abdomen Case #4

• Male European Shorthair cat (7 years)

comes for cleaning teeth

• Pre-anaesthetic lood screening

• In-house analyzer (LasercyteR)

LasercyteR

• Ht 0.24 [0.28-0.47]

• RBC 6.15 [6.0-10.0]

• MCV 39.5 [37.0-55.0]

• MCH 1.03 [0.71-1.07]

• MCHC *** [16.3-22.3]

• Ret% 0.5

• Ret abs 27.8 [3.7-94.1]

• Thromb 229 [156-626]

• Leukocytes 192.1 [6.3-19.6]

• Lymphocytes 116.1 [0.8-4.7]

• Neutrophils 47.5 [3.0-13.4]

• Eosinophils 3.8 [0.3-1.7]

• Monocytes 24.7 [0.0-1.0]

LasercyteR

• Ht 0.24 [0.28-0.47]

• RBC 6.15 [6.0-10.0]

• MCV 39.5 [37.0-55.0]

• MCH 1.03 [0.71-1.07]

• MCHC *** [16.3-22.3]

• Ret% 0.5

• Ret abs 27.8 [3.7-94.1]

• Thromb 229 [156-626]

• Leukocytes 192.1 [6.3-19.6]

• Lymphocytes 116.1 [0.8-4.7]

• Neutrophils 47.5 [3.0-13.4]

• Eosinophils 3.8 [0.3-1.7]

• Monocytes 24.7 [0.0-1.0]

No microscopic evaluation bloodsmear performed

18

Laser flowcytometry

Laser light is scattered by cell

Scattering dependent on different

characteristics

Detectors measure absorbance under

different angles

Laser flowcytometrybed-side instruments (e.g. IDEXX LaserCyte)

Plots Lasercyte in our cat

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Neutropenia

Normal number

leukocytes 40x

Neutrophilia

Use of microscope in assessing neutrophil numbers (20-40 objective)

Case #5

• 13 year old persian cat, female

• Anorexia since 3 days

• Lethargic

• Pale membranes?

• A few ptechiae in oral cavity

• Bloodwork In-House analyzer VetABC Plus

VetABC Plus

• Ht 0.24 [0.28-0.47]

• RBC 6.89 [6.0-10.0]

• MCV 37.0 [37.0-55.0]

• MCH 0.75 [0.71-1.07]

• MCHC 19.5 [16.3-22.3]

• Ret%

• Ret abs [3.7-94.1]

• Thromb 179 [156-626]

• Leukocytes 2.9 [6.3-19.6]

• Lymphocytes 0.4 [0.8-4.7]

• Neutrophils 2.4 [3.0-13.4]

• Eosinophils 0.1 [0.3-1.7]

• Monocytes 0 [0.0-1.0]

VetABC Plus

• Ht 0.24 [0.28-0.47]

• RBC 6.89 [6.0-10.0]

• MCV 37.0 [37.0-55.0]

• MCH 0.75 [0.71-1.07]

• MCHC 19.5 [16.3-22.3]

• Ret%

• Ret abs [3.7-94.1]

• Thromb 179 [156-626]

• Leukocytes 2.9 [6.3-19.6]

• Lymphocytes 0.4 [0.8-4.7]

• Neutrophils 2.4 [3.0-13.4]

• Eosinophils 0.1 [0.3-1.7]

• Monocytes 0 [0.0-1.0]

Remark VetABC Plus analyzer: Plateletclumping; pseudothrombocytopenia

Microscopic evaluation bloodsmear: leukopenia andthrombocytopenia, no platelet aggregates

Impedance/coulter principle(Vet ABC, Medonic)

Sample

Dilution

Lysis Isotonic

WBC Hgb RBC MCV Plt MPV

HctMCHC

(calculated)

20

MIC The MIC flag is

triggered when cells are present in the areaof delineation between thrombocytes and erythrocytes.Possible causes:• Platelet clumps (mainly with cats)• Microcytes

M.W. van Leeuwen, E. Teske:

De hematologische analyzer

VetABC: evaluatie voor gebruik

bij hond en kat. Tijdsch v

Diergeneesk,1999

Case #5 continued

• Thrombocytes not normal but low

• Anaemia, neutropenia and

thrombocytopenia => Pancytopenia

• FelV test: positive

Case #6: Eastern Shorthair, male castrated, 11 years (1612976)

Owner is concerned: cat walked this morning for two meter and than dropped to the floor, lost some urine, but was conscious, lasted a few minutes.

Has not eaten today

Is sleepy the last few weeks

Is current on vaccinations and deworming

Physical Examination

Resp: 30/min

Pulse: 180/min

Temp: 35.7

Muc. Memb: pale

Lnn: normal

Turgor: normal

Respiratory/circulatory/gastrointestinal system: no abn.

DDx Pale membranes

hypovolaemia/shock -

anaemia

- blood loss ±

- hemolysis +

- bone marrow depression +

Initial Diagnostic plan: Ht, reticulocytesBiochemistry

21

Biochemistry• Urea 12.4 mmol/l [5.9-12.5]

• Creatinine 120 µmol/l [71-132]

• Bile acids <8 µmol/l [<8]

• Sodium 147 mmol/l [146-156]

• Glucose 4.7 mmol/l [3.4-5.7]

• Potassium 3.4 mmol/l [3.4-5.2]

• Calcium 2.30 mmol/l [2.3-2.6]

• Phosphate 1.13 mmol/l [0.9-2.0]

• Total protein 90 g/l [54-70]

• Albumin 18 g/l [25-34]

Biochemistry• Urea 12.4 mmol/l [5.9-12.5]

• Creatinine 120 µmol/l [71-132]

• Bile acids <8 µmol/l [<8]

• Sodium 147 mmol/l [146-156]

• Glucose 4.7 mmol/l [3.4-5.7]

• Potassium 3.4 mmol/l [3.4-5.2]

• Calcium 2.30 mmol/l [2.3-2.6]

• Phosphate 1.13 mmol/l [0.9-2.0]

• Total protein 90 g/l [54-70]

• Albumin 18 g/l [25-34]

Hematology – part 1

Hematocrit 0.08 L/L [0.28-0.47]

MCV 51.5 fL [37.0-55.0]

MCH 1.03 fmol [0.71-1.07]

MCHC 20.0 mmol/L [16.3-22.3]

Ret% 5.2%

Ret (abs) 78.5

CHr 1.06 fmol [0.88-1.23]

Platelets 73 x 109 [156-626]

Leukocytes 19.9 x 109 [6.3-19.6]

Neutrophils 18.3 x 109 [3.0-13.4]

Lymphocytes 0.4 x 109 [2.0-7.2]

Blood smear

Hematology – part 1

Hematocrit 0.08 L/L [0.28-0.47]

MCV 51.5 fL [37.0-55.0]

MCH 1.03 fmol [0.71-1.07]

MCHC 20.0 mmol/L [16.3-22.3]

Ret% 5.2%

Ret (abs) 78.5

CHr 1.06 fmol [0.88-1.23]

Platelets 73 x 109 [156-626]

Leukocytes 19.9 x 109 [6.3-19.6]

Neutrophils 18.3 x 109 [3.0-13.4]

Lymphocytes 0.4 x 109 [2.0-7.2]

Blood smear

Peripheral Blood Smear

22

M. Haemofelis (Hemobartonella)

http://www.veteriankey.com

M. haemofelis

Hematology – part 1

Hematocrit 0.08 L/L [0.28-0.47]

MCV 51.5 fL [37.0-55.0]

MCH 1.03 fmol [0.71-1.07]

MCHC 20.0 mmol/L [16.3-22.3]

Ret% 5.2%

Ret (abs) 78.5

CHr 1.06 fmol [0.88-1.23]

PTL 73 x 109 [156-626]

Leukocytes 19.9 x 109 [6.3-19.6]

Neutrophils 18.3 x 109 [3.0-13.4]

Lymphocytes 0.4 x 109 [2.0-7.2]

Blood smear some spherocytes, anisocytosis, polychromasia, Howell Jolly bodies, M Haemofelis

Case management

Blood transfusion for anaemia

Bloodtyping

Lethargy and weakness most likely due to anaemia

Doxycycline for M haemofelis

and…..

Feline hemotropic mycoplasmas

Eperythrozoon felis (1942), Haemobartonella felis (1955), Mycoplasma haemofelis (2001)

Three subtypes:

– Mycoplasma haemofelis (Mhf)

– Candidatus Mycoplasma haemominutum” (cMhm)

– Candidatus Mycoplasma turicensis” (cMtc)

Feline hemotropic mycoplasmas and anemia

Laberke et al (Berl Munich Tierarztl Wochenschr 2010) PCR in 296 cats in Germany (Bavaria)

3 groups: Cats with anaemia, ill cats without anaemia, healthy cats

12.2% positivity (cMhm > Mhf > cMtc)

No significant differences in prevalence between three groups

No association with clinical signs

Jenkins et al (JFMS 2013) PCR in 200 cats in New Zealand

No significant difference in prevalence between anaemic and non-anaemic cats

Vergara et al (Comp Immunol Microbiol Infect Dis 2016) PCR in 384 cats in Chile

No significant difference in prevalence between anaemic and non-anaemic cats, except larger MCV in positive cats

23

Feline hemotropic mycoplasmas and experimental infection

S. Tasker et al: Veterinary Microbiology 2009

10 cats infected with M haemofelis, 3 cats with Cand M haemominutum and 3 cats with Cand M turicensis

PCR, PCV, Coombs (IgM/IgG/Cold (40C)/warm (370C), autoagglutination

Feline hemotropic mycoplasmas and experimental infection

10 cats infected with M haemofelis, 3 cats with Cand M haemominutum and 3 cats with Cand N turicensis

PCR, PCV, Coombs (IgM/IgG/Cold (40C)/warm (370C), auto-agglutination

Only M haemofelis developed macrocytic anaemia

Cold autoantibodies (IgG+IgM) between 8-22 DPI, lasting for 2-4 weeks

Warm autoantibodies (mainly IgG) between 22-29 DPI, lasting for up to 5 weeks

Hematology Case – part 2

Osmotic resistance 10% 0.90 % NaCl [0.58-0.69]

Osmotic resistance 90% 0.75 % NaCl [0.44-0.56]

Coombs:

a-IgM (Fc) (40C) autoagglutination

a-IgG (H+L) (370C) positive (64)

Bloodgroup typing A

Treatment

Bloodtransfusion

Doxycycline 10mg/kg

Prednisolone 2mg/kg

In-house analyzer vs UVDL

• Clinic wanted to compare results In-House analyzer with professional laboratory

• Discrepancy hematocrit was found!

24

Praktijk: vrijdag UVDL: Maandag

StabilityMCV

%flTime

P value

10069,10

106,473,516

105,572,924

0.005111,376,940

0.0021137848

<0.001115,88064

<0.001116,480,472

Hematocrit is calculated by: RBC x MCV

Het UVDL team