Pathogenesis of malaria

INTRODUCTION — Understanding the pathogenesis of malaria requires investigation of mechanisms including parasite invasion, parasite biology, and host defense. Detailed descriptions of the understanding of the underlying molecular biology are provided in the literature [1-3]. The parasite life cycle illustrates the interplay of parasite and host interactions (figure 1). Pathogenesis of Plasmodium falciparum is the area of greatest study, since this species causes the most severe clinical disease (other species include P. ovale [including two subspecies P. o. curtisi and P. o. wallikeri], P. vivax, P. malariae, and P. knowlesi). P. knowlesi malaria can also cause life-threatening illness [4], and, although rare, severe illness (including severe respiratory disease and anemia) and death due to P. vivax have been reported.

Issues related to the pathogenesis of malaria will be reviewed here. Issues related to epidemiology, clinical manifestations, diagnosis, and treatment are discussed in detail separately. (See related topics.)

THE PARASITE

Life cycle — Human malaria occurs by transmission of Plasmodium sporozoites via a bite from an infected female anopheline mosquito (figure 1). The sporozoites travel from the salivary glands of the mosquito through the bloodstream of the host to the liver, where they invade hepatocytes. These cells divide many 1000-fold until mature tissue schizonts are formed, each containing thousands of daughter merozoites. This exoerythrocytic stage is asymptomatic.

The liver schizonts rupture after 6 to 30 days; 98 percent of patients experience liver schizogony by 90 days (there is typically a longer liver phase in species other than P. falciparum). This event releases thousands of merozoites into the bloodstream, where they invade red blood cells (the erythrocytic or asexual stage) in a matter of seconds. P. falciparum may invade any red cell, while P. vivax and P. ovale prefer the younger, slightly larger reticulocytes. The merozoites mature successively from ring forms to trophozoites to mature schizonts (asexual forms) over 24 hours (P. knowlesi), 48 hours (P. vivax, P. ovale, P. falciparum), or 72 hours (P. malariae). Within red blood cells, the parasites digest hemoglobin. As hemoglobin is digested, the toxic metabolite hemozoin (a polarizable crystal) is formed and isolated in the parasite's food vacuole.

The intracellular parasites modify the erythrocyte in several ways. They derive energy from anaerobic glycolysis of glucose to lactic acid, which may contribute to clinical manifestations of hypoglycemia and lactic acidosis [5]. Parasites reduce red cell membrane deformability, resulting in hemolysis and accelerated splenic clearance, which may contribute to anemia. Alterations to uninfected red blood cells, such as the addition of P. falciparum glycosylphosphatidylinositol (GPI) to the membrane, may play a role in increased clearance of uninfected cells and contribute to anemia [6]. (See "Anemia in malaria".)

Ultimately, new daughter merozoites are released from the schizont stage of infected erythrocytes. The remnants of cell membrane and the hemozoin crystal are phagocytized by circulating macrophages, an important stimulus in the activation of the immune cascade [7,8]. In addition, free heme is released into the peripheral blood, an important stimulus for endothelial activation; endothelial cell damage also occurs in some patients [7-9].

Red cell lysis stimulates release of proinflammatory cytokines, including tumor necrosis factor (TNF). TNF suppresses hematopoiesis, which also contributes to the anemia. The liver and spleen enlarge over time; the spleen may become massively enlarged [10]. Thrombocytopenia is caused by a combination of hypersplenism (ie, increased splenic sequestration and decreased platelet survival time) and, in the case of P. falciparum, deposition of platelets adjacent to parasite microvascular sequestration and fibrin thrombi [11-14]. (See 'Coagulation' below.)

Merozoites continue the asexual cycle and infect new red cells, although a few differentiate into male or female gametocytes (sexual forms), which cause no symptoms. These gametocyte-committed parasites leave the bloodstream and develop in the bone marrow over three to four days [15]. Mature male and female gametocytes then circulate in the bloodstream until they are ingested by a blood-feeding female anopheline mosquito. These sexual forms complete their life cycle within the midgut of the Anopheles mosquito (including an ookinete and oocyst stage) with development into sporozoites, which migrate to the salivary glands of the mosquito; from there, they can infect another human through another bite.

In the setting of P. vivax and P. ovale infection, some parasites remain dormant in the liver as hypnozoites and can cause late relapse by reactivating after many months [16]. P. vivax likely has cryptic reservoirs in the spleen and bone marrow [17]. In the setting of P. falciparum and P. malariae infection, hypnozoite parasites do not routinely develop in the liver, but there are rare reports of presumed resurgent P. falciparum infection many years after exposure [18-20]. However, P. malariae can cause attacks even decades after exposure; the mechanism of persistence is unknown. In addition, the chronic infection of P. malariae may result in immune complex formation and deposition leading to renal damage and failure.

The multiple stages of Plasmodium devote a great deal of energy to egress from various cells types in highly regulated processes; these are viable targets for therapeutics [21-23].

Genetic diversity — Surveys of the P. falciparum genome from different geographic regions have demonstrated remarkable genetic diversity, particularly in surface antigens. The parasite genome is much more diverse than the human genome; single nucleotide polymorphisms, insertions/deletions, and microsatellites are very common in particular genes (genes that are under immune selection, for example) [24-26].

Using these polymorphisms as markers in genome-wide association studies allows identification of the source of clinical and experimental phenotypes of the parasites, improving understanding of pathogenesis and potential for development of new therapies. Genetic diversity can also be used as a tool to measure effective population size and the effects of interventions on a given population.

Variation in gene copy number among genes known to be involved in metabolic pathways may influence drug susceptibility [27]. Comprehensive genetic mapping will enable further identification of genes mediating drug resistance as well as potential vaccine targets.

PATHOPHYSIOLOGY

Microvascular disease and sequestration — All species of Plasmodium in human infections are likely cytoadherent to human cell surfaces at some point during the life cycle but, for all species but P. falciparum, this period of sequestration is very short: For P. falciparum, it is more than half of the 48-hour life cycle. Cytoadherence to human cell surfaces is an important component of P. falciparum pathogenesis. As P. falciparum parasites mature from rings to trophozoites within red blood cells, they induce the formation of sticky knobs on the surface of erythrocytes [28,29]. The knobs are composed of a combination of parasite-produced proteins including P. falciparum erythrocyte membrane protein-1 (PfEMP-1, the product of var gene expression and proposed primary cytoadherence factor), KAHRP, PfEMP-2, and RESA as well as human proteins including spectrin, actin, and band 4.1 [30-32]. Each P. falciparum parasite has ≥60 different var genes; of these, one protein product is present in individual parasites [33]. Within the PfEMP-1 molecules, the binding domain cassettes DC8 or DC13 are implicated in severe disease due to their superior binding phenotype for diverse human endothelium [34,35].

The knobs bind to receptors on a variety of cell types in capillaries and venules, including endothelial cells. Notable human receptors include ICAM-1 and ePCR (vascular endothelium), CD36 (on endothelium and platelets), and CSA (in the placenta); a variety of other binding interactions have also been elucidated [36-39]. Endothelial binding leads to sequestration of infected red cells within these small vessels (thereby removing parasites from the peripheral circulation during a prolonged period of the life cycle). This leads to partial blood flow obstruction, endothelial barrier breakdown, and inflammation [28].

Sequestration can be demonstrated in any organ of a patient infected with P. falciparum. The most catastrophic clinical manifestation of sequestration results in cerebral malaria [40]. Renal failure in the setting of malaria may occur in part or as a result of mechanical obstruction by infected erythrocytes; immune-mediated glomerular pathology and fluid loss due to alterations in the renal microcirculation also probably contribute to renal failure [41]. Clearance of infected red cells by macrophages in the spleen through antibody-mediated mechanisms is a crucial control point for preventing severe disease, which is damaged in children with human immunodeficiency virus (HIV) infection [42-45].

Rosetting can occur when infected red cells stick to uninfected red cells and form rosettes that block the microcirculation and contribute to microvascular disease [46,47]. Rosetting is mediated by an interaction between PfEMP-1 within knobs and receptors on the surface of uninfected red cells, such as complement-receptor 1 (CR1) [46,47]. Rosetting occurs in other species, including P. vivax, and has implications for immunopathology and drug resistance [48].

Parasite biomass — Vascular beds harboring sequestered parasites allow accumulation of high levels of parasite biomass in the host. HRP-2 (a secreted P. falciparum antigen expressed on the erythrocyte membrane) can be used as an indirect measure of parasite biomass both in the circulation and sequestered in the microvasculature. The concentration of this antigen has been observed to correlate with severity of clinical disease [49].

Cytokines — The interaction between host endothelium and immune cells with malaria parasites is complex and not fully understood. The "cytokine storm" hypothesis suggests that, in the setting of severe malaria, damaging cytokines and small molecules become unregulated and lead to a systemic inflammatory response syndrome (SIRS)-like state with high circulating levels of tumor necrosis factor (TNF) and nitric oxide. However, evidence of direct correlation between severe malaria and the activity of these markers is limited.

Some markers, such as the acute phase reactant C-reactive protein (CRP), correlate directly with parasitemia. Cytokines TNF, lymphotoxin, interleukin (IL)-6, IL-10, IL-12, IL-18, and macrophage inflammatory protein (MIP)-1 are consistently elevated in the setting of malaria [50]. However, it is not clear whether these precede or follow clinical markers of severe infection.

Molecular evidence for endothelial and tissue damage includes elevated levels of lactate, CK-MB, myoglobin, and angiopoietin-2 as well as increased soluble ligands/receptors (eg, sELAM-1, sICAM-1, sTNF-R1, sTNF-R2, sVCAM-1) [51]. Microparticles (small circulating bodies released from the surface of human cells) are implicated in pathogenesis, host immunity modulation, cell-cell communication, and gametocyte induction with some potential implications for future treatments [52,53].

Coagulation — The initiation of tissue factor production in the coagulation cascade has been proposed as a unifying mechanism of pathogenesis in severe malaria, based on the following observations [11,54-56]:

●Thrombocytopenia is a common feature of severe malaria; it may also be observed in uncomplicated malaria [11,54].

●Activation of the coagulation cascade in the absence of overt bleeding (eg, elevated D-dimer and thrombin-antithrombin complexes with normal prothrombin time and thromboplastin time) is also common [55].

●Autopsies of patients with cerebral malaria frequently demonstrate fibrin microthrombi admixed with platelets in the cerebral vasculature (as well as other organs) [56].

Nitric oxide — Low nitric oxide, low arginine (the precursor of nitric oxide), and elevated arginase activity in peripheral blood have been observed in severe malaria [57]. Metabolic studies have demonstrated that the parasite's arginase enzyme (which converts arginine to ornithine) may contribute to hypoargininemia in severely ill patients, thus shutting down nitric oxide production [58]. Depletion of amino acid precursors for arginine production may also be important [52]. Children with nitric oxide depletion due to intravascular hemolysis in the setting of malaria subsequently develop pulmonary hypertension and myocardial wall stress [57]. Replenishment of nitric oxide via peripheral arginine or inhaled nitric oxide have been suggested as a possible treatment [59-61].

THE HOST

Genetic factors — Several genetic polymorphisms and mutations appear to influence the severity of malaria infection; examples are summarized in the table (table 1) and studies have elucidated addition polymorphisms [62,63].

Hemoglobin and red cell antigens — Hemoglobin and red cell antigens can confer variable protection against malaria. (See "Protection against malaria by variants in red blood cell (RBC) genes".)

●A classic example is the Duffy blood group factor, a red cell antigen necessary for invasion by P. vivax [64]. Absence of the Duffy antigen on red cells (largely observed in individuals from West and sub-Saharan Africa) is protective for P. vivax malaria [64]. However, cases of P. vivax in Duffy-negative individuals have been identified in Brazil and in Kenya, suggesting that P. vivax has evolved alternate red blood cell invasion pathways to invade Duffy-negative red cells [65,66].

●There is strong evidence that sickle cell genetic alterations evolved in part because of the survival advantage against lethal P. falciparum infections [67-69]. Children with HbAS have a significantly lower risk of P. falciparum malaria, lower parasite densities, and lower rates of hospital admissions than children with HbAA [70]. Possible mechanisms are discussed in detail separately. (See "Protection against malaria by variants in red blood cell (RBC) genes".)

The potential protective effect of sickle hemoglobin against malaria may be augmented in malaria-endemic areas; individuals outside endemic areas may have a lesser degree of protection. In a family living in the United States in which two children had sickle cell anemia and three had sickle cell trait, travel to an endemic region without chemoprophylaxis led to hemolytic crisis in three of the children [71]. Thalassemia may indirectly protect against P. falciparum infection by mediating increased susceptibility to nonlethal P. vivax, particularly in young children [72,73].

●Red blood cells in individuals with thalassemia appear to be susceptible to P. falciparum invasion but are associated with significantly reduced parasite multiplication [74,75]. This may be due to the variable degree of persistence of hemoglobin F, which is relatively resistant to hemoglobin digestion by malarial hemoglobinases [73,76,77]. (See "Pathophysiology of thalassemia", section on 'Reduced malaria risk'.)

●Ovalocytosis in Southeast Asia appears to confer protection against malarial infection. Possible mechanisms include diminished invasion, poor intraerythrocytic growth, or diminished cytoadherence of infected erythrocytes [78,79].

●Hereditary elliptocytosis appears to confer protection against malaria, although malaria infection in the setting of this condition has been described [80]. (See "Protection against malaria by variants in red blood cell (RBC) genes", section on 'Hereditary elliptocytosis (HE)'.)

●The haptoglobin (Hp) genotype determines the efficiency of hemoglobin clearance after malaria-induced hemolysis. A particular polymorphism of the haptoglobin genotype (Hp2/2) has been associated with a reduction in the number of clinical malarial episodes; this was illustrated in a study of 312 children in Kenya [81].

●Pyruvate kinase deficiency appears to be protective against infection and replication of P. falciparum in human erythrocytes. Therefore, mutant pyruvate kinase alleles may confer protection against malaria in endemic areas [82].

Tumor necrosis factor — Polymorphisms in tumor necrosis factor (TNF) genes appear to influence the severity of P. falciparum infection [83,84]. This was illustrated in a study of approximately 1000 Gambian children; a sevenfold increased risk of severe neurological sequelae or death from cerebral malaria was observed among those who were homozygous for a polymorphism in the promoter region of the TNF gene (TNF2 allele). Severe anemia was associated with a different TNF allele, suggesting that different genetic factors affect susceptibility to these two disease manifestations [83].

Immunity — Individuals living in endemic areas appear to develop partial immunity to clinical episodes of malaria following repeated infections; the degree of protective immunity appears to be proportional to transmission intensity and increases with age. Individuals in highly endemic areas (eg, sub-Saharan Africa) acquire nearly complete protection from clinical disease by early adulthood [85-87]. Individuals in low transmission areas (eg, Southeast Asia) remain at risk for clinical disease and fatal disease into adulthood; these individuals are referred to as "semi-immune."

Individuals not living in endemic areas (eg, travelers) infected with malaria form a detectable antibody response (which can be measured by enzyme-linked immunosorbent assay [ELISA]), although this response is not protective against the initial infection of malaria and may serve only as a marker of past exposure [87].

Immunity in pregnancy is also important. The risk of parasitemia during the first years of life is higher among children born to multigravid women than primigravid women [88,89]. The immunologic basis for this observation is not fully understood.

Humoral response — The humoral immune response to malaria appears to correlate with severity of clinical infection, with progression in maturation of the humoral response in the setting of ongoing parasite antigen stimulation. Elevated levels of immunoglobulin (Ig)G4, IgE, and IgM are associated with severe disease in individuals with ≤5 previous clinical episodes of malaria, while elevated levels of IgG (IgG, IgG1, IgG2, and IgG3) are associated with mild disease in individuals with >5 previous clinical episodes [90,91].

These observations suggest that persistence of humoral immunity requires ongoing parasite antigen stimulation. In addition, individuals who leave endemic areas appear to lose some humoral protection; these individuals are "semi-immune," and their protection is lessened when they return to endemic areas after prolonged periods without parasite antigen stimulation. (See "Prevention of malaria infection in travelers".)

Cellular response — The ability to mount a robust interferon-gamma response (predominantly through CD56+ gamma T cells) has been associated with protection against high parasitemia [92]. Phagocytosis of hemozoin or trophozoites impairs the ability of monocytes and macrophages to mount oxidative burst, kill ingested bacteria, present antigens correctly, and mature to functioning dendritic cells [7]. These cells produce TNF and other proinflammatory cytokines and release peroxidation derivatives of polyunsaturated fatty acids [7]. In addition, these cells have increase in activity and release of MMP-9, which is correlated with TNF- and interleukin (IL)-1 gamma production and leads to disruption of the basal lamina of endothelial cells [7,8]. Neutrophils, which release toxic granules as well as neutrophil extracellular traps, may play a role in malaria pathogenesis; the inhibition of these cells in the setting of malaria infection likely predisposes to secondary infection [93].

SUMMARY

●Understanding the pathogenesis of malaria requires investigation of mechanisms including parasite invasion, parasite biology, and host defense. The parasite life cycle illustrates the interplay of parasite and host interactions (figure 1). (See 'Introduction' above.)

●The Plasmodium life cycle consists of an exoerythrocytic (asymptomatic) stage and erythrocytic (symptomatic) stage. Plasmodium sporozoites are transmitted by the bite of an infected female anopheline mosquito. The sporozoites invade hepatocytes, which divide until schizonts are formed containing thousands of daughter merozoites. These rupture and release merozoites into the bloodstream, where they invade red blood cells. (See 'Life cycle' above.)

●Intracellular parasites alter the red cell, digesting hemoglobin to form hemozoin and making the membrane less deformable, resulting in hemolysis or splenic clearance. The merozoites invade the red cell and mature to ring forms, trophozoites, and schizonts. Schizonts rupture and release new daughter merozoites. This release can result in proinflammatory cytokines response including tumor necrosis factor. Most released merozoites infect new red cells; a few differentiate into gametocytes, which circulate until they are ingested by a mosquito to continue the transmission cycle. (See 'Life cycle' above and 'Cytokines' above.)

●Endothelial binding of infected red cells leads to sequestration of infected red cells within small vessels (thereby removing parasites from the peripheral circulation during a prolonged period of the life cycle). This can lead to partial blood flow obstruction, endothelial barrier breakdown, and inflammation. Mechanisms of microvascular disease include formation of sticky knobs on the cell surface and rosetting (eg, adherence of infected red cells to uninfected cells, forming rosettes that clog the microcirculation). (See 'Microvascular disease and sequestration' above.)

●Several human genetic polymorphisms and mutations have been observed to influence the severity of malaria infection, particularly hemoglobin and red cell antigens. (See 'Genetic factors' above and "Protection against malaria by variants in red blood cell (RBC) genes" and "Anemia in malaria".)

●Individuals living in endemic areas develop partial immunity to malaria following repeated infections; they develop protection from severe disease and can be asymptomatic despite infection. The degree of protective immunity appears to be proportional to transmission intensity. The cellular immune response consists of a variety of cytokines including interferon-gamma and tumor necrosis factor. (See 'Immunity' above.)