A n n a l s o p C l i n i c a l a n d L a b o r a t o r y S c i e n c e , Vol. 4, No. S Copyright © 1974, Institute for Clinical Science

R ed Cell Indices in Thalassem ia Minor

HENRY G. SCHRIEVER, M.D.

Department of Pathology, John F, Kennedy Community Hospital,Edison, NJ 08817

ABSTRACT

Red cell size and hemoglobin content, as measured by the Coulter Counter Model S, are consistently and rather uniformly decreased in thalassemia minor. Red cell count, hemoglobin and hematocrit are more variable and may even be normal. The finding of mean corpuscular volume (M CV) 53 to 71 p? and mean corpuscular hemoglobin (M C H ) 17 to 25 p g should raise the suspicion of thalassemia minor. The M CV/RBC quotient is regularly under 13 in thalassemia minor and over 13 in non-thalessemic microcytosis except for some overlap in women. A presumptive diagnosis can then be made by the finding of target cells and basophilic stippling of red cells on the peripheral blood smear. Demonstration of elevated A2 hemoglobin level clinches the diagnosis. ( RBC = red blood cells.)

Beta thalassemia minor is an hereditary hypochromic microcytic anemia, probably a mutation favored by selection over many generations in Mediterranean peoples to provide protection against the malarial par­asite.2’6’10 While thalassemia minor is harm­less enough, the homozygous state of the gene produces a severe anemia, thalas­semia major or Cooley’s anemia, which is usually fatal before adulthood.7 Most pa­tients with thalassemia minor are either not anemic or are only mildly anemic. Most are quite unaware of their condition. It is important to establish a correct diagnosis to avoid unnecessary iron treatment and to counsel patients about the possibility of thalassemia major in their offspring.

This report presents the diagnostic blood findings in a series of thalassemia minor patients. A simple stepwise procedure for

making the diagnosis is outlined which can be used for inexpensive mass screening for the condition. Complete blood counts were done on 93 patients with thalassemia minor on the Coulter Counter Model S .* Pe­ripheral blood smears were stained with Wright-Giemsa and examined in the usual manner. Quantitative hemoglobin A2 levels were done by a modification of the poly­acrylamide gel electrophoresis method of Bierman and Zettner.1

The results of the automated blood counts are shown in table I. All of the patients with thalassemia showed signifi­cant basophilic stippling of red cells and target cells on smear, features which were not at all prominent in cases of iron de­ficiency anemia with red cells indices in the thalassémie range. All patients with

* Coulter Electronics, Hialeah, F L 33010.

3 4 0 SCH RIE VER

T A B L E IP e r i p h e r a l B l o o d i n 93 P a t i e n t s w i t h T h a l a s s e m i a M i n o r

M e a n ± S t a n d a r d D e v i a t i o n

NHemoglobin

g p e r d l

Red C e ll Count

1 0 6 p e r cmmH em atocrit

P ercen tMCV*p3

MCH*p g

MCHC P erce n t

Men 29 1 2 .6 0 ± 1 .3 6 5 .9 8 ± 0 .4 8 37 .6 ± 3 ,8 4 6 2 .8 + 3 .8 8 2 0 .7 ± 1 .5 7 3 3 . 1 3 ± 0 .8 6

Women 45 1 1 . 5 9 ± 1 .0 8 5 .3 8 + 0 .5 3 34 .8 9 ± 3 .1 6 6 4 .2 2 + 3 .5 1 2 1 .3 3 ± 1 .4 0 3 3 .1 8 + 0 .9 8

C h i l d r e n 19 1 1 . 2 9 + 1 . 1 5 5 .6 3 + 0 .4 7 3 3 , 8 4 ± 3 .2 6 5 9 .6 8 ± 3 .3 0 1 9 . 7 ± 1 . 1 0 33 ,2 6 ± 1 .0 7

N o r m a l M 1 4 - 18

i—! 1r- 42 - 52 82 - 92 27 - 3 1 32 - 36

F 1 2 - 16 4 .2 - 5 .4 37 - 47 8 1 - 94 27 - 3 1 32 - 36

*M ean c o r p u s c u l a r v o l u m e tM e an c o r p u s c u l a r h e m o g lo b i n 'VMean c o r p u s c u l a r h e m o g lo b in c o n c e n t r a t i o n

in men greater than in children (p < 0.05); MCV in women greater than in children ( p < 0.01); and MCH in women greater than in children (p < 0.002). The other differences were not statistically significant. Mean corpuscular hemoglobin concentra­tion (M C H C ), a value which is sometimes decreased in iron deficiency anemia, was normal in all but five cases. The overall thalassemic range for MCV was 53 to 71 p? and for mean corpuscular hemoglobin MCH from 17.5 to 25.4 pg.

England and Fraser3 recently developed a mathematical “discriminant function” (D F ) to differentiate thalassemia minor from iron deficiency:

D F = MCV-RBC-(5 X H b) - 3.4

MCV is expressed in /xz, RBC in millions per cmm and Hb in grams per dl. They found the D F regularly negative in thalas­semia minor and positive in iron deficiency. In table II is shown the D F in thalassemia minor and non-thalassemic microcytosis. Mentzer5 proposed an even simpler for­mula to differentiate the two conditions: M CV/RBC; he found this quotient to be usually over 13 in iron deficiency. In table III is shown this value in thalassemia and non-thalassemic microcytosis. The M CV/ RBC quotient worked for men and chil­

thalassemia minor showed elevation of he­moglobin A2 in the 4.2 to 8.9 percent range (normal up to 4.1 percent). The t-test11 showed the following differences to be statistically significant: hemoglobin and hematocrit in men greater than those in women (p < 0.01); RBC in men greater than in women (p < 0.002); m ens hemo­globin and hematocrit greater than chil­dren s (p < 0.002); RBC in men greater than in children (p < 0.05); MCV in men greater than in children (p < 0.01); MCH

T A B L E II

D i s c r i m i n a n t F u n c t i o n (D F) i n T h a l a s s e m i a

T r a i t a n d N o n - t h a l a s s e m i c M i c r o c y t o s i s

D F = M C V - R B C - (5 X Hb) - 3.4

DF-f-(Range)

DF-(Range)

Non-thalassemia * (MCV under 80)

19(2.1-60.2)

1(-13)

Thalassemia trait

Men 1(2.2) (-0.7

28-- 22.5)

Women 17(2.6-8.6) (0.0

28--12.8)

Children 2(1.2-2.0) (-0.1

17-- 24.1)

*Mean corpuscular volume

RED C E L L INDICES IN TH A LA SSEM IA M INOR 341

dren, but the thalassemic women ranged higher than 13 (up to 15.3), overlapping with the range for non-thalassemic micro- cytosis both in men and women. The high values for women are due partly to their significantly lower RBC and partly to their tendency toward higher MCV values. Co­existing iron deficiency was not a factor in the one thalassemic woman with M CV/ RBC of 15.3; her serum iron was normal.

Screening for thalassemia minor could proceed as follows:

1. The MCV and MCH are measured, looking for values in the thalassemic range, and the M CV/RBC value is calculated.

2. Where indices and/or the M CV/RBC value are suspicious, the peripheral blood smear is examined for target cells and basophilic stippling. In a routine laboratory situation critical review of the blood film may be nec­essary.

3. Confirmation of the diagnosis is made by precise measurement of the A2 hemoglobin fraction by a quantitative

M e a n C o r p u s c u l a r V o l u m e P e r R e d B lo o d C e l l s ( M C V / R B C ) i n T h a l a s s e m i a a n d

N o n -t h a l a s s e m ic M ic r o c y t o s is

TABLE III

MCV/RBCf u n d er 13

(R an ge o f V a lu e s )

MCV/RBCf o v e r 13

(R an ge o f V a lu e s )

N o n - t h a l a s s e m i a

* (M CV u n d e r 8 0 )0 20

( 1 3 . 0 - 1 9 . 5 )

T h a l a s s e m i a t r a i t

M e n 28( 0 . 6 - 1 2 . 7 )

1( 1 4 . 1 )

W om en 34( 9 . 4 - 1 3 . 0 )

1 1( 1 3 . 1 - 1 5 . 3 )

C h i l d r e n 1 9( 8 . 7 - 1 2 . 8 )

0

* M e a n c o r p u s c u l a r v o l u m et M e a n c o r p u s c u l a r v o l u m e p e r r e d b l o o d c e l l s

technique. In rare cases of thalas­semia minor, A2 hemoglobin may be normal, and hemoglobin F elevated.4

In studying families known to have thal­assemia, A2 hemoglobin determination on

F i g u r e 1 . Peripheral blood smear in beta thal­assemia minor showing target cells and baso­philic stippling. When red cell indices are in the thalassemic range, these findings may be enough for a presump­tive diagnosis of thalas­semia minor. (X 1,000)

3 4 2 SCH RIEV ER

each member is hardly necessary, the di­agnosis being obvious from the cell indices and blood film. Additional laboratory tests, such as iron and iron binding capacity, osmotic fragility, bone marrow examination and the usual hemoglobin electrophoresis done for hemoglobinopathy, are super­fluous in most cases. Alpha thalassemia trait is also prevalent in Mediterranean and Oriental peoples8-10 and may be present when the M CV/RBC value and the periph­eral blood smear suggest thalassemia al­though hemoglobins A2 and F are normal.9 Erythrocyte morphology may be normal in alpha thalassemia trait, however.10

R eferences1. B i e r m a n , A. H. a n d Z e t t n e r , A.: A simple

electrophoretic method for the quantitative determination of hemoglobin A2. Amer. J. Clin. Path. 37 :139-142, 1967.

2. C h e r n o f f , A.: The distribution of the thal­assemia gene: a historical review. Blood 14: 897-912, 1959.

3. E n g l a n d , J . M. a n d F r a s e r , P.: Differentia­tion of iron deficiency from thalassemia trait by routine blood count. Lancet 1:449-452, 1973.

4. M a r k s , P. A.: Thalassemia syndromes. New Eng. J. Med. 275:1363-1369, 1966.

5. M e n t z e r , W. C.: Differentiation of iron de­ficiency from thalassemia trait. Lancet 1:882,1973.

6. M o t u l s k y , A. G.: Current concepts of the genetics of the thalassemias. Cold Spring Har­bor Symposium on Quantitative Biology 29: 399-413, 1964.

7. N a t h a n , D. G.: Mild thalassemia—a guide to therapy. New Eng. J. Med. 281:1363—1364,1969.

8. N a t h a n , D. G.: Thalassemia. New Eng. J. Med. 286:586-594, 1972.

9. P e a r s o n , H. A., O ’ B r i e n , R . T., a n d M c ­

I n t o s h , S.: Screening for thalassemia trait by electronic measurement of mean corpuscular volume. New Eng. J . Med. 288:351-354,1973.

10. R u c k n a g l e , D. L .: On the geographical dis­tribution and ethnic origin of thalassemia. New Zea. Med. J. Hema. Suppl. 65:826-831, 1966.

11. Statistical Analysis. Olivetti Corporation o f America, New York, pp. 188-193, 1968.

Plan to attend the 25th Anniversary M eeting of the

ASSOCIATION OF CLINICAL SCIENTISTS

in Philadelphia, PA on November 8, 9 and 10, 1974

Applied Sem inar on the

Laboratory Diagnosis o f Skeletal, M uscular and

Nervous Disorders