The increased clinical state of malaria infection may be
due to poor nutritional
status most especially as a
result of micronutrients deficiency. Micronutrients play vital role both in combating anaemia
and other adverse effects of malaria infection in humans and animals in
developing resistance against the disease. This research was aimed at establishing the effect
of malaria parasite on neutrophil-lymphocyte ratio, platelet-lymphocyte ratio and some essential
plasma electrolytes (calcium, magnesium and iron) as well as vitamins (vitamin B12 and
folate) of children. Ethical
approval was gotten from ministry of Health, Benin City, Edo State. Test group was recruited from the
paediatric ward of Central Hospital, Benin City. A questionnaire, written in English was
administered to the parents of the subjects to fill. An informed consent was given to the parents or
guardian of the children before
sample was collected Blood sample was collected from two hundred (n = 200) children within the ages of
l – 10 years as test group who are positive for malaria parasites while the
control group were one hundred (100) children above l0yrs of age who do not have malaria
parasites. Thick blood film was used to ascertain the level of parasitaemia using giemsa stain,
vitamin B12 was done using electroluminescence technique, folic acid
was analysis was done using ELISA technique, while iron, magnesium and calcium was spectrophotometrically analyzed. Results gotten in this
research shows that PCV decreased significantly compared to the control group,
(p<0.05). White blood cell increased (p<0.05) compared to
the control, Plasma calcium, magnesium and iron decreased significantly;
there was decrease in folate and vitamin B12. Micronutrients are not
only
necessary in the regeneration of heamolyzed red cells during malaria infection,
but also served as antioxidants hence protecting the red cells against damage by
malaria toxins. It is therefore of tremendous importance to assess micronutrients
status of children with malaria.
TABLE
OF CONTENTS
TITLE
PAGE. i
CERTIFICATION.. ii
DEDICATION.. iii
ACKNOWLEDGEMENT. iv
TABLE OF
CONTENTS. v
LIST OF TABLES. x
LIST OF
FIGURES. xi
ABSTRACT. xii
CHAPTER ONE. 1
INTRODUCTION.. 1
1.1 Background of Study. 1
1.2 Justification of Study. 4
1.3 Aim of Study. 4
1.4 Specific Objective. 5
1.5 Research Design. 5
1.6 Ethical consideration. 5
1.7 Research Hypothesis 5
1.8 Questionnaire/informed consent 6
1.9 Sample size. 6
CHAPTER TWO.. 7
2.0 LITERATURE REVIEW.. 7
2.1 Malaria. 7
2.2 Epidemiology. 7
2.3 Malaria parasites 10
2.3.1 Life cycle of Malaria Parasite. 11
2.3.2 Signs and Symptoms of Malaria in
Children. 13
2.3.3 Consequences of Malaria in Children. 13
2.3.4 Severe Malaria Anaemia in Children. 14
2.3.5 Laboratory Diagnosis of Malaria. 14
2.4 Micronutrients 15
2.4.1 Functions of Micronutrients 17
2.4.2 Micronutrients Deficiency in children. 18
2.4.3 The Prevalence of Micronutrient
Deficiency in Diet Plans 19
2.5 Vitamin B12 (Cobalamin) 20
2.5.1 Structure of Vitamin B12 20
2.5.2 Metabolism of Vitamin B12;
Absorption. 21
2.5.3 Mechanism of Absorption of Vitamin B12 22
2.5.4 Transportation, Storage and Excretion
of Vitamin B12 23
2.5.5 Functions of Vitamin B12 25
2.5.6 Deficiency of Vitamin B12 25
2.5.7 Laboratory Assessment status of Vitamin
B12 26
2.6 Folic Acid (Folate) 26
2.6.1 Structure of Folic acid. 26
2.6.2 Formation of Folic acid (F.H4) 27
2.6.3 Biosynthesis of Folic acid. 29
2.6.4 Functions of Folic acid. 30
2.6.5 Deficiency of Folic acid. 30
2.6.6 Laboratory Assessment Status of Folic
acid. 31
2.7 Iron (Fe) 31
2.7.1 Essential Iron. 32
2.7.2 Storage Iron. 32
2.7.3 Absorption of Iron and Factors
Regulating Absorption. 33
2.7.4 Iron Transport and Utilization. 35
2.7.5 Iron Requirements 36
2.7.6 Iron Deficiency (I.D) 36
2.7.7 Diagnosis of Iron deficiency. 37
2.7.8 Iron toxicity. 38
2.8 Calcium (Ca) 38
2.8.1 Dietary sources and Absorption of Ca. 39
2.8.2 Mechanism of Ca Absorption. 39
2.8.3 Factors Affecting Ca Absorption. 39
2.8.4 Regulation of Ca Concentration. 41
2.8.5 Functions of Ca. 42
2.8.6 Hypercalcaemia and Hypocalcaemia. 43
2.8.7 Measurement of total Ca. 43
2.9 Magnesium.. 44
2.9.1 Chemistry. 45
2.9.2 Dietary Sources 46
2.9.3 Absorption, Transport, Metabolism and
Excretion of Magnesium.. 47
2.9.4 Functions Of Magnesium.. 50
2.9.5 Hypermagnesaemia. 51
2.9.5.1Causes
of Hypermagnesaemia. 51
2.9.6 Hypomagnesaemia. 52
2.10 Effects
of Malaria parasite on Micronutrients in Children. 53
2.11 Iron
Levels in Malaria parasite Infection. 53
2.12 Calcium
Levels in Malaria Parasite Infection. 54
2.13 Magnesium
Levels in Malaria Parasite Infections 55
2.14 Folate
Levels in Malaria Parasite Infection. 55
2.15 Vitamin
B12 Levels in Malaria Parasite Infection. 57
CHAPTER THREE. 58
3.0 MATERIAL AND METHODS. 58
3.1 Study Area. 58
3.2 Study Population. 58
3.3 Inclusion criteria. 58
3.4 Exclusion criteria. 58
3.5 Control group. 58
3.6 Sample Collection. 58
3.7 Laboratory Analysis 59
3.7.1 Procedure For Malaria Parasite Test 59
3.7.2 Assay Of Serum Calcium Concentration. 59
3.8 Quality Control 61
3.9 Analysis For Vitamim B12 And Folic
Acid. 61
3.9.1 Electroluminescence (ECL) For
Measurement of Vitamin B12. 61
3.9.2 Methodology/Steps 62
3.9.3 Enzyme-linked Immunosorbent Assay Kit
For Folic Acid (FA) 63
3.9.4 Reagents and Quantity. 64
3.9.5 Methodology/Steps 64
3.9.6 Assay Of Serum Magnesium Concentration. 66
3.10 Procedure For Serum Iron. 68
3.11 Statistical Analysis 69
CHAPTER FOUR.. 70
4.0 RESULTS. 70
CHAPTER FIVE. 77
DISCUSSION.. 77
CONCLUSION
AND RECOMMENDATION.. 80
REFERENCES. 81
Table 1: Food sources of Magnesium
(Saris et al., 2000) 47
Table 2: Distribution of magnesium in
the adult human. 49
Table 3: Procedure/protocol table for
calcium estimation. 60
Table 4: Procedure/Protocol table for
magnesium estimation. 67
Table 5: Age, sex, and malaria load
of study and control subjects 70
Table 6: Comparison between the PCV,
RBC, HgB conc., red cell indices, total and differential leucocyte counts, NLR,
and PLR of the study and control groups. 73
Table 7: Comparing the calcium,
magnesium, iron, vit. B12, and folic acid levels between the malaria Positive
subjects and Malaria negative subjects 75
Figure 1: Malaria cases (per 100,000)
by Country Worldwide. 9
Figure 2: Life Cycle of Malaria
Parasite (Ayodotun and
Olugbenga, 2012). 12
Figure 3: Structure of Vitamin B12
(Chatterjea and Shinde, 2012). 21
Figure 4: Mechanism of Absorption of
Vit. B12 (Chatterjea and Shinde, 24
Figure 5: Structure of Folacin. 27
Figure 6: Steps involved in the
Formation of F.H4. 28
Figure 7: Structure of F.H4. 29
Figure 8: Absorption of Iron. 35
Figure 9: Electroluminescence method,
competitive principle for measuring vitamin B12. 63
Figure 10: showing the ages of the
study and control groups 71
Figure 11: showing the sex of the
study and control groups 71
Figure 12: Comparison between the
PCV, RBC, HgB conc., and red cell indices, of the study and control groups. 74
Figure 13: Comparing the calcium,
magnesium, iron, Vit. B12, and folic acid levels between the malaria Positive
subjects and Malaria negative subjects 76
Malaria is a prevalent
disease in tropical and subtropical areas of Africa. It is estimated that 1-3
million deaths occurs worldwide, mostly involving children under the age of
5years (Gouado et al., 2007). Malaria
is a major public health challenge in Nigeria and it accounts for more cases
and deaths than any other country in the world (Olasehinde et al., 2010). This disease is often linked to changes
in climate, poverty, malnutrition and the double resistance of the malaria
parasite to usual anti-malaria drugs and insecticides (Müller and Garenne,
1999). Infection by malaria can cause serious health problems and this often
leads to death especially in children (Gouado et al., 2007). The disease is caused by malaria parasites
(Plasmodium species) which are transmitted by the female anopheles mosquito
(vector). There are today more than 25 named plasmodium species which infect
primates. Four of the species are human parasites; P. falciparum, P. vivax, P. malaria and P. ovale (Trampuz et al., 2003). Epidemiological studies have demonstrated that P. falciparum is the most dangerous specie as it is responsible for most of the
deaths caused by malaria (Greenwood et al.,
2005). The malaria parasite is transmitted when an individual is bitten by
infected female anopheles mosquito (Ochei and Kolhatkar,
2008). The main symptom of uncomplicated malaria in children is fever. Older
children may present with headache, backache, chills, myalgia and fatigue (Olasehinde
et al., 2010). Severe anaemia may exist alone or in combination
with other complications particularly cerebral malaria and respiratory distress
in which it portends worse prognosis (WHO, 2004). Consequences of severe
malaria include coma and death if untreated, young children are especially
vulnerable (Anemana et al., 2004).
Laboratory diagnosis of malaria could be made by detection of parasite in blood]
or by serological techniques (Ochei and Kolhatkar, 2008).
Micronutrients
are trace elements that are required in small quantities to ensure normal
metabolism, growth and physical well‐being. Some studies relating micronutrient status and malaria
infection reported low plasma levels of certain micronutrient in acute malaria
infection (Alonso, 2004). Iron and beta carotene which are reported to have
modulatory effect on the pathogenesis of malaria, have been observed to be
deficient in acute plasmodium falciparum infection (Lavender, 1993; Shankar
and Prasad 1998;; Beard, 2001; Caulfield et al., 2004). The
levels of micronutrients in children are of particular interest since adequate
intake is of great importance for the well being, proper development, and
functioning of the body starting from fetal life and throughout childhood. Micronutrients
comprise of vitamins and minerals. Examples of vitamins are vitamin A, pro-vitamin
A (Beta‐carotene),
vitamin B1,
vitamin B2,
vitamin B6,
vitamin B12, biotin, vitamin C, vitamin E, vitamin D, vitamin K
,folic acid, niacin and pantothenic acid while minerals include the trace
elements such as iron, copper, iodine, manganese, selenium and zinc together
with the macro elements calcium, magnesium, potassium and sodium (Asaolu and Igbaakin, 2009; Crook, 2012).
Micronutrients have been implicated to play important roles in immunity and
physiologic functions. For instance, Calcium is an important nutrient that
plays a major role in bone and teeth formation, impulse transmission, catalytic
activation among others (Nordin, 1997). Iron plays an important role in the
production of heamoglobin, oxygenation of red blood cells and lymphocytes. It
improves the function of enzymes in protein metabolism and enhances the
function of calcium and copper (Asaolu
and Igbaakin, 2009). Vitamin B12 is involved in the
maturation of red blood cells. The folic acid
coenzymes are specifically concerned with metabolic reactions involving the
transfer and utilization of the one carbon moiety (Crook, 2012). Micronutrients are
found in small quantities within the body and they are obtained from a wide
variety of foods. No single food
contains all of the micronutrients we need and, therefore, a balanced and
varied diet is necessary for an adequate intake. Micronutrients deficiency is
more frequent amongst children in developing countries (Gibson and Ferguson,
1998). These deficiencies may contribute to an increased risk of parasitic
infection such as malaria (Mahomed, 2000).
Malaria
has been a major challenge to the world’s population especially in Africa and
indeed Nigeria. It has been implicated in increased rate of morbidity and mortality
among children (Sachs and Malaney, 2002). Research
has shown that malaria causes 0.5 – 3.0 million
deaths each year and that 75% of these deaths occur in African children under
the age of 5years (Greenwood et al.,
2005).The increased clinical state of malaria infection may be due to
poor nutritional status more especially as a result of micronutrients
deficiency (Gouado et al., 2007). Micronutrients play vital role both
in combating anaemia and other adverse effects of malaria infection in humans
and animals in developing resistance against the disease. Micronutrients are
not only necessary in the regeneration of heamolyzed red cells during malaria
infection, but also served as antioxidants hence protecting the red cells
against damage by malaria toxins (Jain, 2006). It is therefore
of tremendous importance to assess micronutrients status of children with
malaria.
The aim of this study is
to assess the levels of micronutrients in children with malaria infection in Paediatric
Ward, Central Hospital, Benin city.
1. To
evaluate the levels of vitamin B12,
folic acid, iron, magnesium, and calcium in malaria infected children.
2. To
correlate the micronutrient levels with the severity of malaria infection.
3. and
to compare the results with the control
group (non-infected children).
This
is a case study designed to assess micronutrients status of malaria infected
children and then compare findings with non-infected children in Benin City.
Ethical approval was sought and approved
by the ethical committee of Central Hospital, Benin City, Edo State.
H1: malaria parasites affects
calcium, magnesium, iron ,vitamin B12,folic acid in children
infected with malaria.
H0 malaria
parasites does not affect calcium, magnesium, iron ,vitamin B12,folic
acid in children infected with malaria.
A
questionnaire, written in English was administered to the parents of the
subjects to fill. An informed consent was given to the parents or guardian of
the children before sample was collected.
The
sample size (N) was calculated using
prevalence from previous studies
The
sample (Olasehinde et al., 2010). size for this study will be obtained
using the formula described by Dean et
al., (1995).
N = Z2P (1 – P)
D2
N
= required
sample size
Z = confidence
level at 95% (standard value of 1.96)
P = estimated
prevalence of intestinal parasites of pupils (84.7%)
D = margin
of error at 5% (standard value = 0.05)
N=1.962
X 0.847(1-0.847)
0.052
3.8416 X
0.847(0.153)
___________________
0.0025
N=199 minimum sample sizes
Therefore
a minimum of 200 test samples and 100 control will be used for this research.
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