Apicomplexa: Malaria
3.1 Malaria
Over half of the world’s population is at risk from catching malaria, a disease that results from infection with protozoan parasites of the genus Plasmodium.
At the turn of the 20th century, Sir Ronald Ross discovered that malaria re- sulted from bites of infected Anopheline mosquitoes, but before that it was be- lieved that malaria was the result of unhygienic conditions, or ‘mal-air’ (bad air). There are five species of Plasmodium that infect humans:
r
Plasmodium falciparum and Plasmodium vivax account for the majority of morbidity and mortality associated with malaria.r
The distribution and prevalence of Plasmodium ovale and Plasmodium malariae infections are both much lower.r
Plasmodium knowlesi (formerly known as a primate malaria) is now recog- nised as a significant pathogen of humans in South-east Asia.Malaria is currently endemic in 109 countries in four continents and, of the 500 million cases of malaria estimated to occur annually, approximately one million result in death. Most of the fatalities are in children under the age of five years old and pregnant women.
The degree to which populations become exposed to malaria can vary, depend- ing on the transmission rate. In areas where malaria is holoendemic (with most of the population infected to some degree), transmission can be high, but a non-sterile immunity that accommodates asymptomatic infection builds with age. In other areas, where rain patterns dictate fluctuations in the mosquito population, malaria can be both seasonal and unstable.
3.1.1 The life cycle of malaria
The life cycle of Plasmodium consists of several different developmental stages (both intracellular and extracellular), controlled by a genome consisting of over
Immunity to Parasitic Infection, First Edition. Edited by Tracey J. Lamb.
C2012 John Wiley & Sons, Ltd. Published 2012 by John Wiley & Sons, Ltd.
5,000 genes. The parasite largely replicates as a haploid organism, with the only diploid stages occurring in the mosquito (ookinete/oocyst stage).
3.1.1.1 Sporozoites in the skin
Malaria infection is initiated upon deposition of sporozoites into the avascular tissue of the skin from the salivary glands of a female mosquito as it probes for a blood meal (Figure 3.1). Within one minute, the sporozoites become highly motile, traverse the capillary wall and enter the blood stream. However, some sporozoites can remain motile in the skin for several hours, while others enter the lymphatic system and can be found in the draining lymph nodes, where the host mounts an immune response.
MOSQUITO MIDGUT
10-14 days
Diploid ookinete (~10µm) Sporozoites
(10x1µm)
LIVER 7-10 days
Oocyst (10-20µm)
Mosquito feeds ingesting gametocytes Mosquito feeds depositing sporozoites
Gametocytes
ņ Ņ
Arrested hypnozoites (P. vivax / P. ovale)
Merozoites (1-2 µm)
BLOODSTREAM Asexual erythrocytic cycle
Ring
Trophozoite
Schizont
Schizont rupture
24-72 hours depending on species
Haemozoin crystal
MAMMALIAN HOST
Figure 3.1 The life cycle of malaria.Malaria sporozoites are deposited in the vascular beds of the skin by a mosquito bite;
these then actively traverse the endothelium, migrating to the liver via the bloodstream. They traverse several hepatocytes before developing in to a large exoerythrocytic form (LEF). InP. vivaxandP. ovaleinfections, some sporozoites invade hepatocytes but undergo arrested development to form hypnozoites, which are largely resistant to drug treatment and are responsible for relapsing malaria infections. Merozoites develop inside the LEFs and burst out of the hepatocyte to invade erythrocytes. During maturation in erythrocytes, the malaria parasites export proteins to the surface of the erythrocyte, remodelling it and enabling removal from the circulation via sequestration to the endothelium of a number of organs in the body. Some erythrocytes invaded by merozoites do not continue cycling, instead developing to become transmissible stages known as gametocytes. When taken up by mosquitoes, male and female gametocytes mate in the midgut of the mosquito and undergo several stages of development before becoming sporozoites, which migrate to the salivary glands and are deposited upon the next feed.
3.1.1.2 Liver-stage malaria
Once in the liver, sporozoites glide along the sinusoidal epithelium traversing several Kupffer cells (resident liver macrophages) before invading a final hepa- tocyte, in which a parasitophorous vacuole (PV) forms. The infected hepatocyte then grows into a large exoerythrocytic form (LEF), which eventually gives rise to between 10,000 and 20,000 merozoites over a 7–10 day time period.
The liver-stage of the life cycle is not associated with notable disease in malaria infection, but allows the parasite to multiply. Relapsing malaria infections caused by P. vivax and P. ovale arise from arrested liver-stage parasites known as hypnozoites, which are generally resistant to anti-malarial drugs.
3.1.1.3 Asexual erythrocytic cycle
Once merozoites burst from hepatocytes they invade red blood cells (RBC) and enter into the asexual erythrocytic cycle. This stage of malaria is associated with most of the pathology in malaria infection, and the length of each replication cycle differs depending on the species of malaria parasite (Figure 3.1).
Invasion of new RBCs by merozoites involves secretion of proteases from struc- tures found at the apical end of the merozoites called micronemes, rhoptries and dense granules. One of the major surface proteins of merozoites is a 200 kDa protein called merozoite surface protein (MSP)-1. This protein is essential for asexual cycling in RBC stages; it is proteolytically processed in a number of stages after it reaches the merozoite surface, a process necessary for invasion.
P. vivax requires the presence of a glycoprotein on the RBC surface (known as Duffy antigen) to attach. This species is not a major contributor to the malaria burden across Sub-Saharan Africa, where most of the population are of the Duffy-negative blood type, making their RBCs refractory to invasion by P. vivax.
Once inside the RBC, the parasite becomes once again encased by a PV mem- brane but it is able to remodel the surface of the RBC. Parasite proteins are syn- thesised, secreted from the parasite and exported over the PV membrane to the RBC surface. The passage of the proteins across the PVM and into the RBC cytosol is mediated by a complex of proteins forming a translocation channel (or translocon) known as Plasmodium translocon of exported proteins (PTEX).
Proteins that traffic to the surface of the infected RBC (e.g. P. falciparum ery- throcyte membrane protein (Pf EMP)-1) tend to contain motifs known as Plas- modium export elements (PEXEL motifs) to facilitate this process, although some exceptions exist.
RBCs do not have nuclei and are essentially metabolically inactive cells. Repli- cating parasites obtain the amino acids they require by digesting haemoglobin.
Additionally, RBC modification by the parasite renders the RBC more perme- able to essential anions, sugars, amino acids and organic cations from the blood plasma, in a process termed the ‘new permeation pathway’. At schizogony, once infected RBCs burst, between 10–32 merozoites are released, and these invade fresh RBCs to being a new erythrocytic cycle.
3.1.1.4 Transmission back to mosquitoes
A small proportion of iRBC differentiate in to transmissible male and female gametocytes, but the exact molecular cues leading to the development of male and female gametocytes are unknown. Once inside the mosquito midgut, the temperature shift and pH change induces gametogenesis and fertilisation lead- ing, to the formation of motile diploid ookinetes that leave the blood meal bolus and traverse the midgut epithelium to become sessile oocysts. Over 10–14 days, sporozoites develop within the oocyst via mitosis, and these escape via an enzy- matic process into the mosquito body cavity. The sporozoites circulate via the haemolymph and attach onto the basal lamina of the mosquito salivary glands, ready for introduction into the next host.
3.1.2 Mouse models of malaria
Much of the current knowledge on immune responses to malaria infection have been derived from a combination of observations in human malaria infections, in vitro modelling of immune responses to malaria, and mechanistic investi- gations using animal models of malaria. The mouse has a well-characterised immune system that is easy to manipulate and, although no one mouse model replicates all of the symptoms of human malaria infections, there are many similarities in the immune responses and pathology observed.
The phenotype of infection that results in mouse models of malaria depends on the parasite species, as well as the strain of mouse being infected. Cerebral malaria caused by P. falciparum is often modelled by Plasmodium berghei in- fections of mice (on a susceptible C57BL/6 background) or Plasmodium yoelii infections, whereas malarial anaemia and memory immune responses are of- ten studied using non-lethal infections such as Plasmodium chabaudi.