PREVALENCE AND SEVERITY OF HYPOGLYCAEMIA AND LACTIC ACIDOSIS IN CHILDREN DIAGNOSED WITH PLASMODIUM FALCIPARUM MALARIA

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ABSTRACT

The prevalence and severity of hypoglycaemia and lactic acidosis in Nigerian children diagnosed with Plasmodium falciparum malaria were determined in 100 outpatient children aged 3-144 months (12 years). The children were grouped into 2 categories: 3-59 month old and 60-144 month old. The results obtained indicated that out of the 100 children recruited into this study, seventy-five (75%) were infected while twenty-five (25%) were uninfected with Plasmodium falciparum malaria. On the basis of age group, higher incidence of malaria was recorded in children under 5 years of age with prevalence rate of 85.3%, while those above 5 years had low prevalence rate of 14.7%. The mean blood glucose concentration of malaria-infected children below 5 years (3.80 ± 0.73 mmol/l) was lower than that of malaria-infected children above 5 years (4.21 ± 1.34 mmol/l); however, the difference was not significant (p>0.05). Comparatively, the mean glucose concentrations of the corresponding uninfected subjects were 4.10 ± 0.87 and 4.26 ± 0.51 mmol/l respectively. The mean blood lactate concentration of children below 5 years of age (2.59 ± 1.63 mmol/l ) was significantly (p<0.05) higher than those above 5 years (2.30 ± 1.75 mmol/l). The mean values for both groups were also above the normal range of 1.0 – 2.0 mmol/l while the mean haemoglobin concentration of malaria-infected children below 5 years (16.11 ± 2.24 g/dl) was slightly lower than that of malaria-infected children above 5 years (16.36 ± 2.64g/dl) though not significant (p> 0.05). The prevalence rates of 14.7% were recorded for both hypoglycaemia and lactic acidosis in malaria-infected subjects while 16.0% was recorded for anaemia. There was no significant correlation between blood lactate concentration and blood glucose concentration (r= 0.032, p=0.751) but there was significant positive correlation between haemoglobin level and glucose concentration (r=0.401, p=0.0001). The results suggest that the risk of hypoglycaemia, lactic acidosis and anaemia is higher in younger children, particularly among those below five years of age and also confirmed the knowledge that malaria is a major cause of hospital visits by children.

TABLE OF CONTENTS

                                                                                                                                           PAGE

Title Page                                                                                                  i         

Certification                                                                                                 ii

Dedication                                                                                                      iii

Acknowledgements                                                                                        iv

Abstract                                                                                                             v

Table of Contents                                                                                     vi

List of Figures                                                                                       x

List of Tables                                                                                                 xi

List of Abbreviations                                                                                   xii

CHAPTER ONE: INTRODUCTION

1.1        Malaria                                                                                                  2

1.1.1     World malaria report                                                                                 3

1.1.2     Malaria in children                                                                                       5

1.1.3     Malaria parasite life cycle                                                                      5

1.1.3.1 Sporogony within the mosquitoes                                                          5

1.1.3.2 Schizogony in the human host                                                    6

1.1.3.3 Pre-erythrocytic phase-schizogony in the liver                                      6

1.1.3.4 Erythrocytic schizogony-centre stage in red cells                       7

1.1.4     Pathogenic basis of malaria                                                             9

1.1.5     Pathophysiology of severe malaria in children                      11

1.1.6     Cytokine-associated neutrophil extracellular traps and antinuclear

antibodies in Plasmodium falciparum                                               12

1.2        Biochemistry of Plasmodium falciparum                                13 

1.2.1     Detoxification of heme and reactive oxygen intermediates      17

1.2.2     Biochemistry and molecular biology of malaria parasite:

             pyrimidine biosynthetic pathway                                              18

1.2.3     Complication of Plasmodium falciparum malaria                                 20

1.2.4      Prevalence and management of Plasmodium falciparum malaria       among infants and children                                                             22

1.3        Hypoglycaemia in childhood malaria                                        22

1.3.1     Sublingual sugar for hypoglycaemia in children with severe malaria                            24

1.4        Lactic acidosis in childhood malaria                                            24

1.4.1    Lactate levels in severe malarial anaemia                       25    

1.5       Transport of lactate and pyruvate in Plasmodium falciparum malaria                          26        

1.6       Anaemia in childhood malaria                                                         27

1.6.1    Severity of anaemia in children diagnosed with Plasmodium falciparum    malaria                                                                                                                                     28

1.7       Typhoid and malaria co-infection                                                 28

1.8       Aim and objectives of the study                                                        29

1.8.1    Aim of the study                                                                                     29  

1.8.2    Specific objectives of the study                29                                                

CHAPTER TWO: MATERIALS AND METHODS

2.1       Materials                                                                                          30

2.1.1    Subjects and location                                                                            30

2.1.2    Instruments/Equipment                                                                    30

2.1.2.1 Accutrend plus meter                                                                       30

2.1.2.2 Crista haemoglobinometer                                                             31

2.1.3    Reagent kit/ Test strips                                                                         31

2.1.3.1 Malaria diagnostic rapid test kit                                                       31

2.1.3.2 Glucose reagent strip                                                            31

2.1.3.3 Lactate reagent strip                                                                               31

2.2       Methods                                                                                                 31

2.2.1    Preparation of 70% (v/v) ethanol                                                        31

2.2.2    Experimental design                                                                     32

2.2.3    Malaria diagnostic test                                                                      32

2.2.4    Biochemical parameters determined                                         33

2.2.4.1 Determination of blood glucose concentration                 33

2.2.4.2 Determination of blood lactate concentration                                     34

2.2.5    Haematological parameter determined                                                35

2.2.5.1 Estimation of haemoglobin concentration                                35

2.3       Statistical analysis                                                                  35

CHAPTER THREE: RESULTS

3.1   Prevalence of Plasmodium falciparum malaria according to age of subjects                 36

3.2   Blood glucose levels in malaria-infected and uninfected subjects       37

3.3   Blood lactate levels in malaria-infected and uninfected subjects       38

3.4   Haemoglobin levels in malaria-infected and uninfected subjects       39

3.5   Variation of glucose concentration with age of malaria-infected       subjects                                                                                                    40

3.6   Variation of lactate concentration with age of malaria-infected        subjects                                                                                       41

3.7   Variation of haemoglobin concentration with age of malaria-infected          subjects                                                                                       43

3.8   Effect of Plasmodium falciparum parasite load on the blood glucose         concentration of subjects                                                          44

3.9   Effect of Plasmodium falciparum parasite load on the blood lactate         concentration of subjects                                                     46

3.10 Effect of Plasmodium falciparum parasite load on haemoglobin        concentration of subjects                                                                      47

3.11 Comparison of glucose concentration of malaria-infected and         uninfected subjects                                                                   48

3.12 Comparison of lactate concentration of malaria-infected and         uninfected subjects                                                             49

3.13 Comparison of haemoglobin concentration of malaria-infected and         uninfected subjects                                                                                     50

3.14 Correlations matrix                                                                         51

CHAPTER FOUR: DISCUSSION

4.1    Discussion                                                                                          53

4.2   Conclusion                                                                                                  56

4.3   Suggestions for further studies                                                          56

REFERENCES                                                                               57     

APPENDICES                                                                                                76

LIST OF FIGURES

Fig 1: World malaria burden (World Malaria Report, 2011)                          4

Fig. 2: Ingestion of host cytoplasm                                                          15

Fig. 3: Variation of glucose concentration with age of malaria-infected             subjects                                                                 40

Fig. 4: Variation of lactate concentration with age of malaria-infected             subjects                                                                     42

Fig. 5: Variation of haemoglobin concentration with age of malaria-infected             subjects                                                                               43

Fig. 6: Effect of Plasmodium falciparum parasite load on the blood glucose

           concentration of subjects                                                      45

Fig. 7: Effect of Plasmodium falciparum parasite load on the blood lactate

           concentration of  subjects                                                             46

Fig. 8: Effect of Plasmodium falciparum parasite load on haemoglobin

           concentration of  subjects                                                            47

Fig. 9: Comparison of glucose concentration of malaria-infected and

            uninfected subjects                                                                       48

Fig. 10: Comparison of lactate concentration of malaria-infected and

            uninfected subjects                                                                    49

Fig. 11: Comparison of haemoglobin concentration of malaria-infected and

             uninfected subjects                                                                      50

LIST OF TABLES

Table 1: Indicators of severe malaria and poor prognosis                     21

Table 2: Prevalence of Plasmodium falciparum malaria according to age of subjects           36

Table 3: Blood glucose concentration (BGC) in malaria-infected and uninfected   subjects                                                                                   37

Table 4: Blood lactate concentration (BLC) in malaria-infected and uninfected               subjects                                                               38

Table 5: Haemoglobin concentration (HC) in malaria-infected and uninfected   subjects                                                                           39

Table 6: Correlations matrix                                               52

LIST OF ABBREVIATIONS

µl:                                           Microlitre

ABC:                                      ATP-binding cassette

ANOVA:                               Analysis of Variance

ATC:                                       Aspartate transcarbamylase

ATP:                                       Adenosine triphosphate

BGC:                           Blood glucose concentration

BLC:                           Blood lactate concentration

CPS :                          Carbamyl phosphate synthase

CRP:                           C-reactive protein

DBL:                          Duffy binding-like

de novo:                      From the beginning

DHO:                         Dihydroorotase

DHOD:                      Dihydroorotate dehydrogenase

DNA:                         Deoxyribonucleic acid

DPAP:                       Dipeptidyl aminopeptidase

DRC:                         Democratic Republic of the Congo

Fe2+:                          Ferrous ion

Fe3+:                          Ferric ion

GSH:                         Reduced glutathione

H+:                            Hydrogen ion

HC:                           Haemoglobin concentration

HCl:                          Hydrochloric acid

HCM:                        Hz-containing monocytes

HCN:                        Hz-containing neutrophils

HCO3:                                  Bicarbonate

HRP:                         Histidine-rich protein

Hz:                            Haemozoin

IFN-y:                       Interferon gamma

IgG:                           Immunoglobulin G

IgM:                              Immunoglobulin M

IL:                                 Interleukin

LED:                             Light-emitting diode

MCTs:                          Monocarboxylate transporters

NAD+:                          Nicotinamide adenine dinucleotide

NADH:                         Reduced form of nicotinamide adenine dinucleotide

NETs:                            Neutrophil extracellular traps

OMPDC:                      Orotidine 5’-monophosphate decarboxylase

OPRT:                          Orotate phosphoribosyltransferase  

pCMBS:                       p chloromercuribenzenesulphonate

Pfmdr-1:                       Plasmodium falciparum multidrug resistance protein/gene

pHi:                              Intracellular pH

pLDH:                         Plasmodium lactate dehydrogenase

ppm:                             Parasite plasma membrane

PVM:                           Parasitophorous vacuole membrane

RBC:                            Red blood cell

RNA:                           Ribonucleic acid

ROI:                             Reactive oxygen intermediates

SD:                              Standard deviation

SEARO/WPRO:         South-East Asia and Western Pacific Regional Office

SOD:                           Superoxide dismutase

SPPS:                          Statistical product and service solutions

TNF-α:                        Tumor necrosis factor alpha

TRAP:                        Thrombospondin-related anonymous protein

UMP:                          Uridine 5’ monophosphate

WHO/AFRO:             World Health Organization Regional Office for Africa

CHAPTER ONE

                                             INTRODUCTION

Plasmodium falciparum is the most common cause of severe and life-threatening malaria, which causes over 2 million deaths every year (Bruneel et al., 2003; Njuguna and Newton, 2004). In Africa, a vast majority of these deaths occur in children under five years of age (WHO, 2012). Lactic acidosis complicates 35% of severe childhood malaria (Krishna et al., 1994) and hypoglycaemia is present in 20% of children with cerebral malaria (Newton and Krishna, 1998). Both acidosis and hypoglycaemia commonly coexist but each is considered separately as a cause of fatality in children and adults due to severe complicated malaria. Hypoglycaemia is known to be an independent risk factor for death in both severe malaria (Gray et al., 1985; Molyneux et al., 1989) and other severe childhood infections in the tropics (Kawo et al., 1990). Despite its importance, its pathogenesis is not well understood (English et al., 1998). Hypoglycaemia is associated with a poor prognosis in severe malaria (krishna et al, 1994).

In African children with malaria, impairment in hepatic gluconeogenesis in the presence of adequate levels of precursors (glycerol) has been considered the most likely mechanism (White et al., 1987). Irreversible coma may quickly develop if the condition is not effectively treated. Hyperlactataemia is often associated with a poor outcome in severe malaria in African children (Krishna et al, 1994). The pathophysiology of metabolic acidosis is complex. The direct contribution of P. falciparum to the final lactate concentration, through anaerobic glycolysis in the parasite itself, is likely to be small (Vander et al., 1990). More significantly, an inadequate supply of oxygen to tissues may follow from severe anaemia and provoke a metabolic shift within host cells to anaerobic glucose metabolism and increased lactic acid production. In addition, the flow of blood through the microcirculation may be impeded by adherence of infected erythrocytes to the endothelium of post-capillary venules and/or increased rigidity of uninfected cells (Dondrop et al., 1997). Lactate may not in itself be sufficient to cause acidaemia but the inhibition of oxidative metabolism in the context of an ongoing inflammatory response will cause protons (H+) to accumulate and eventually lead to metabolic acidosis (English et al.,1997). These pathophysiological pathways suggest that the syndrome of lactic acidosis may be associated with the total parasite burden during acute infection.

Acute malaria is estimated to cause 225 million cases of ill health per year, resulting in over one million deaths per year, most of which occur in sub-Saharan Africa (World Malaria Report, 2010; Murray et al., 2012). Malaria is particularly virulent among children, constituting one of the principal causes of child morbidity as well as mortality in sub-Saharan Africa (WHO, 2000). Exposure to the malaria parasite not only results in bouts of high fevers among children, but also increases the risk of malnutrition and anaemia among children under five (Ehrhardt et al., 2006).

  1. Malaria
PREVALENCE AND SEVERITY OF HYPOGLYCAEMIA AND LACTIC ACIDOSIS IN CHILDREN DIAGNOSED WITH PLASMODIUM FALCIPARUM MALARIA

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