Predicting iron and folate deficiency anaemias from standard blood testing: the

mechanism and implications for clinical medicine and public health in developing
countries
Alan E Dugdale
1
1
Department of Paediatrics and Child Health, School of edicine, !niversity of
"ueensland, " #$$%, Australia
Corresponding author&
Alan E Dugdale: a&dugdale'acenet&net&au
(eceived April )$, )$$%* Accepted +ctober ,, )$$%&
-his is an +pen Access article distributed under the terms of the Creative Commons
Attribution .icense /http:00creativecommons&org0licenses0by0)&$1, 2hich permits
unrestricted use, distribution, and reproduction in any medium, provided the original
2or3 is properly cited&
+ther Sections4
o Abstract
o 5ac3ground
o ethods and results
o Discussion
o Abbreviations
o Competing interests
o Authors6 contributions
o (eferences
Abstract
5ac3ground
Developing countries have high prevalence of diseases, but facilities to diagnose and treat
them are limited& 7e must use available resources in 2ays not needed 2here there are
sophisticated e8uipment and trained staff& Anaemia is common* iron deficiency affects
health and productivity* folate deficiency in pregnant 2omen causes foetal abnormalities&
9e2 developing countries can measure serum folate or ferritin, but standard automated
blood analyses are 2idely available and can help predict folate and iron deficiency& -he
(D7:C;< /coefficient of variation of the red cell 2idth1 measures the variability in the
si=e of red blood cells /(5C1 in routine automated analysis of blood cells, but is seldom
reported& .evels of (D7:C;< and haemoglobin /Hb1 can predict iron deficiency
anaemia&
ethod and results
> have 2ritten a computer model based on the standard mechanism for blood formation
and destruction& -his sho2s that before anaemia develops and during recovery, there are
both normal and abnormal (5C /small in iron deficiency and large in folate deficiency1
in the circulation& -he model calculates the abnormality in the (D7:C;< in standard
automated blood analyses& >n early iron deficiency and during recovery the full blood
count sho2s the Hb near the lo2er limit of normal, a lo2 C; and a high (D7:C;<&
A similar pattern, but 2ith a higher C;, develops in folate deficiency& 9olate
deficiency is often brief and may not cause anaemia& -he high (D7:C;< may persist
for three months&
Conclusion
-his long footprint could be medically useful for detecting folate deficiency and so
limiting foetal damage in individuals and communities& 9e2 clinicians or public health
2or3ers 3no2 about (D7:C;<& Standard blood reports for clinical use should include
the (D7:C;< and note the possible significance of abnormal values&
+ther Sections4
o Abstract
o 5ac3ground
o ethods and results
o Discussion
o Abbreviations
o Competing interests
o Authors6 contributions
o (eferences
5ac3ground
A recent paper ?1@ has confirmed the findings of earlier authors ?),A@ that 2e can predict
the onset of iron deficiency anaemia using output from automated blood analy=ers& -hese
papers sho2 that the blood changes parallel the lo2 levels of iron stores& -he main
indicators are haemoglobin level /Hb1 near the lo2er limit of normal and a high level of
anisocytosis measured by the coefficient of variation /<1 of the red cell distribution
2idth /(D7:C;<1& Bo mechanism for this finding 2as proposed& > describe a computer
simulation model that follo2s the formation and destruction of red blood cells /(5C1&
7hen standard assumptions about red cell formation and destruction are used, and 2ith
an ade8uate supply of iron, the output of the model corresponds to the findings in normal
blood& 7hen iron is deficient the hitherto uneCplained changes appear& -he model also
eCplains 2hy people 2ith macrocytic anaemia may have normal folate level even though
the changes in the blood are due to folate deficiency ?#@&
-he mechanism of the model is as follo2s& 7hen normal bone marro2 has ade8uate ra2
materials and hormonal stimulus, it produces enough normal (5C to maintain circulating
Hb levels& -he mean values for circulating (5C are mean cell volume /C;1 ,$ D 1$
and (D7:C; 1)<& 7hen there is insufficient iron for normal haemopoiesis, the marro2
produces microcytes 2ith C; %$ D E& After the start of altered haemopoiesis, the
circulation contains a miCture of normal cells and microcytes, so the (D7:C;<
increases rapidly, 2ell before the overall levels of C; and Hb drop belo2 the normal
range& 7hen folate is lac3ing, the marro2 produces macrocytes* the miCture of
macrocytes and normal (5C raises the (D7:C;< before the C; and Hb become
abnormal&
-he computer model
-he model is a computer programme that follo2s the changes in the (5C contained in
one cubic millimetre of blood& -he model is based on a matriC, 1)$ columns 2ide* each
column contains the number and properties of (5C formed in a single day& -he model
runs 2ith intervals of one /simulated1 day& -he ne2 cells formed are entered into Column
1 of the matriC& At each iteration /day1, the cohort of cells is moved one column to the
right: Column 1 :F Column ), Column 1$ :F Column 11, and so on& >t is assumed that the
(5C in Column 1)$ have been destroyed and lost to the circulation& >n this simple
version of the model, a life span of 1)$ days is assumed& > also ta3e the lo2er level for
normal of Hb as 11$ g0l /the 7orld Health +rganisation6s lo2er limit for normal Hb for
adult 2omen1 and the upper limit of normal (D7:C;< as 1G<&
-he input data are /a1 the number of cells formed, /b1 the mean cell volume, /c1 the
standard deviation of the mean cell volume, /d1 the mean cell haemoglobin, /e1 the
number of days during 2hich these conditions apply& >t is assumed that the type of
haemopoiesis s2itches from one form to another, e&g& normal to iron:deficient, 2ithin one
day, but this is not critical for the 2or3ing of the model& -his simple version of the model
also assumes that all (5Cs have a lifespan of 1)$ days&
At the start of the run the matriC is empty& -o populate the matriC, the model is run for
1)$ days 2ith normal values for each of the input parameters& At any iteration, the
characteristics of the (5C contained in one cubic millimetre of blood can be sho2n& -he
output values are (5C count per cubic mm, haemoglobin g0l, mean cell volume /C;1,
mean cell haemoglobin /CH1, mean cell haemoglobin concentration /<1 /CHC<1
and the red:cell distribution 2idth /(D7:C;<1& +nce the matriC is populated, the type
of haemopoiesis can be changed and the effect on the (5C parameters sho2n&
+ther Sections4
o Abstract
o 5ac3ground
o ethods and results
o Discussion
o Abbreviations
o Competing interests
o Authors6 contributions
o (eferences
ethods and results
>ron deficiency
-able 1 sho2s the effects of iron deficiency on the full blood count& -he model is first
run for 1)$ days 2ith normal haemopoiesis to populate the matriC /(5C0day #$$$$,
C; ,$, SD of C; 1$, CH A$1& 9ollo2ing this, the model runs for A$ days /Days $
H A$1 2ith normal haemopoiesis, then for 1G$ days /Days A$ H 1I$1 2ith iron deficient
haemopoiesis /(5C0day A$$$$, C; %$, SD of C; E, CH 1I1&
Table 1
-he changes in standard haematological findings 2ith iron:deficient
haemopoiesis&
-he C;, CH and (D7:C;< in the blood volume are initially normal& 5y the end of
A$ days of abnormal haemopoiesis, the Hb and the C; have decreased but remain
2ithin normal limits& Ho2ever, the (D7:C;< becomes abnormally high, going from
11&)< to 1I&A<, because of the miCture of circulating microcytes and normocytes& After
G$ days of iron deficiency the Hb is 111, still 2ithin the normal range, but the (D7:C;
< has risen further to )1&A<& -he Hb continues to fall and the (D7:C;< continues to
rise until all the normal cells formed before the iron:deficient haemopoiesis have been
removed from the circulation& After this, there is a uniform population of iron deficient
(5C, the Hb level stabili=es and the (D7:C;< returns to normal levels& -he critical
finding is that the (D7:C;< becomes abnormal 2hile the Hb and C; are still 2ithin
normal range& -his eCplains the findings ?),A@ that a high (D7:C;< predicts later iron
deficiency anaemia& 7hen iron therapy is given and normal haemopoiesis returns /not
sho2n here1, there 2ill again be both normal and iron:deficient (5C in the circulation so
the (D7:C;< 2ill again rise to abnormal levels until the microcytes formed during the
period of iron deficiency reach the end of their lifespan&
9olate deficiency
-he body has eCtensive stores of iron, so iron deficiency is li3ely to be a long:term event
producing the typical anaemia& 9olate is a 2ater:soluble vitamin& 5ody stores are
relatively small and labile so temporary reduction of dietary inta3e can produce short:
term /less than 1 month1 folate deficiency, 2hich is relieved by a fe2 meals of folate:
containing foods& ;egetable and fruit are the usual sources of folate* in developing
countries, seasonal shortages often occur* in 2estern countries, poor people may forgo
these foods 2hen less cash is available from 2elfare or other payments& !sually this
2ould have little effect on health, but if the 2oman is in the first trimester of pregnancy,
even a temporary deficiency of folate could produce severe and permanent effects on the
foetus& -he model sho2s that short:term deficiency produces characteristic effects on the
(5C parameters&
-able ) sho2s the changes associated 2ith short:term folate deficiency& As in -able 1,
the model is populated 2ith normal (5C and then run for another A$ days 2ith normal
haempoiesis& 9rom Day 1$ to Day #$, haemopoiesis shifts from normal to the macrocytic
pattern of folate deficiency& 9rom Day #$ on2ards, the folate deficiency has been
corrected and haemopoiesis returns to the normal mode& During this short period of folate
deficiency, the Hb drops and the C; rises, but both remain 2ithin normal range&
Ho2ever, the (D7:C;< rapidly rises beyond the normal range& >t remains high for
more than 1$$ days after the end of the folate deficiency, that is until the cohort of
macrocytes has left the circulation 1)$ days after the folate level has returned to normal&
Table 2
-he changes in standard haematological findings 2ith temporary folate:
deficient haemopoiesis&
+ther Sections4
o Abstract
o 5ac3ground
o ethods and results
o Discussion
o Abbreviations
o Competing interests
o Authors6 contributions
o (eferences
Discussion
-his model puts numerical values to the 2ell:3no2n process of (5C formation and
destruction& >n so doing it sho2s the cause for hitherto uneCplained observations ?1:A@
and predicts other clinical applications for routine blood analysis& -he model sho2s that
in iron deficiency, 2hich could be due to inade8uate iron stores or unavailability of iron
resulting from acute infection, there is an early and prolonged rise in the (D7:C;<
before the other parameters indicate anaemia& -he (D7:C;< remains high until the
blood is populated entirely by hypochromic cells& 7hen iron is given and the
haemopoiesis returns to normal, there is again an increase in the (D7:C;< /not sho2n
in -able 11 for as long as there is a miCed population of normal and microcytic (5C in
the circulation&
-he model sho2s similar findings in short:term folate deficiency& -he C; increases
and Hb decreases, but these remain 2ithin normal limits, 2hile the (D7:C;< rises
rapidly beyond the normal range& 9olate levels can 8uic3ly return to normal 2hen folate
is fed, but the haematological effects remain for several months after the return of normal
haemopoiesis until the macrocytes leave the circulation& -his eCplains the lac3 of
correlation bet2een serum folate levels and macrocytic anemia ?#,G@1&
-his method of detecting early iron and folate deficiencies is designed for use in
countries 2here iron deficiency is very common H up to G$< in 2omen of child:bearing
age H folate deficiency much less common and other causes, 2ith the eCception of
malaria and thalassaemia in some countries, uncommon& -he model suggests that
measures of C; and (D7:C;< have t2o functions: first, to detect the possibility of a
problem* second, to determine the nature of the problem& !chida ?)@ reported a sensitivity
of EE&1< for iron deficiency anaemia, #,&)< for iron deficiency anaemia plus latent iron
deficiency, and specificity ,$&%<& >n thalassaemia minor, the (D7:C;< is more li3ely
to be normal ?%@, but Jreen ?E@ stated that the discrimination is not good and other
parameters that are measured by automated analysers but not reported, such as the cell
haemoglobin distribution, may be better& >n the anaemia of thalassaemia, the
haemoglobin level is lo2 but the (D7:C;< is usually normal ?I@& >n the anaemia if
chronic illness the (D7:C;< is often 2ithin the normal range ?,@ and in malaria the
(D7:C;< is lo2 ?1$@, but the best discriminator is a lo2 platelet count ?11@&
-his method cannot distinguish bet2een folate and ;itamin 51) deficiencies, but 51)
deficiency is much less common in most developing counties& 7hen more than one of
these essential nutrients is lo2, then the measurements of Hb, C; and (D7:C;<
reflect only the effects of the limiting nutrient& >t is most unli3ely that more than one of
these 2ill be a limiting nutrient at the same time& 9or eCample, if iron and folate levels
are both lo2 but iron is limiting nutrient, then the (5C 2ill be small and hypochromic,
2ith a lo2 C; and Hb and a high (D7:C;<& >f iron is given to correct the iron
deficiency, then there 2ill be a partial response until haemopoiesis is limited by the lo2
folate& At this time the C; and (D7:C;< 2ill sho2 the folate deficiency&
-he sensitivity and specificity of this method of predicting iron deficiency cannot be
calculated from the computer model but are given in the papers cited& -he sensitivity and
specificity for folate deficiency cannot be determined& >f there is long:term folate
deficiency, serum folate levels 2ill correspond 2ith the haematological changes, but
(obinson and ladonovic ?#@ note that serum folate levels may be normal even in the
presence of macrocytic anaemia& 9or short:term folate deficiencies that do not lead to
anaemia but may cause foetal damage, the changes in C; and (D7:C;< remain long
after the folate level has returned to normal& -here may be no other method for detecting
recent folate deficiency and hence no gold standard to calculate sensitivity and
specificity&
-his is a very simple model designed to sho2 the causes of hitherto uneCplained changes
in the C; and (D7:C;< and their clinical significance& 9or simplicity, > have
assumed that (5C formation is either normal or abnormal /iron deficient or folate
deficient1 although the model changes little if 2e assume a gradual transition& > have
made no attempt to include the effects of other factors such as erythropoietin on blood
formation& -his method of detecting deficiencies in iron and folate does not supplant
standard measures of serum iron and folate, but rather provides a useful tool in regions
2here anaemia is prevalent and resources limited&
-echnical description of the model
-he model is 2ritten in D+S 5AS>C language and 2ill run on any PC:based computer& A
compiled version and the source code are both available& Details and code of the
computer model are available from the author&
Abbreviations
Hb Haemoglobin level /g0l1
(D7:C;< Coefficient of variation of the red cell 2idth /<1
C; ean (ed cell ;olume /fl1
CHC< ean Cell Haemoglobin Concentration /<1
Ht Haematocrit
CH ean Cell Haemoglobin /pg1
Competing interests
-he author/s1 declare that they have no competing interests&
Authors6 contributions
-he author elucidated the mechanism described, designed and 2rote the computer
program and also 2rote the paper&
Figure 1
Changes in Hb and (D7:C;< 2ith iron deficiency&
Figure 2
Changes in Hb and (D7:C;< 2ith folate deficiency&
Ac3no2ledgements
> than3 Dr -ommaso Cavalli:Sfor=a, Butrition Adviser, 7H+ (egional +ffice, anila
for his continuing advice and help and s Bisha Khan, Chief Dietician, Jovernment of
9iLi for her help&