jackie's flashcards malaria

Understand the global epidemiology of malaria infections

• Still close to the #1 infectious cause of child mortality
• Access to available interventions remains poor
• Drug and insecticide resistance rife
• Poor predictors of severe malaria
• No widely available vaccine and need new drugs

3 stages of Malaria

1. mosquito stage ( sexual reproduction )

2. liver stage ( asexual reproduction

3. Blood stage ( asexual reproduction, major amplification stage )

Describe the life cycle of the malaria parasite

1. sexual stage in the female anopheles mosquito ( 1-2 weeks )

2. injected sporozoites (a motile spore-like stage in the life cycle of parasites ) enter the hepatocytes via skin (Hepatocytes are the primary functional cells of the liver)

3. asexual liver stage ( 1-2 w)

4. asexual blood stage cycle is relatively synchronous ( 2-3 days )

5. disease occurs a week to a month after infection

6. gametocytes ( the sexual precursor cells of malaria ) form in the blood and are taken up by a feeding mosquito

describe the disease pathogenesis of the malaria parasite

• fever chills anaemia

• most deaths ( 95%) are caused by P falciparum ( cerebral ( coma ), severe anaemia, placental malaria )

• P.vivax results in severe morbidity but low mortality

• relapses, hypnozoites in liver

Why isn't it identifiable in the blood?

parasitic RBC stick to blood vessel walls ( cytoadherence ) removing itself from the cirulation to mature so when you take a blood sample of someone with a malria you often don't see anything becasue its not in the cirulating blood. Parastie generally lie in skeletal muscle or fat tissue, kidney or brain ( sequester to different tissue )
WHY?: because if they ciruclated, the spleen would spot the irregular RBC

what is the PfEMP1/var genes

The PfEMP1 proteins and var genes play a central role in how Plasmodium falciparum (the parasite that causes the most severe form of malaria) evades the immune system and causes disease.

What are Var genes

encodes the different variants of PfEMP1.
Only one var gene is expressed at a time, in a process called mutually exclusive expression.
The parasite can switch between different var genes during infection → changing the PfEMP1 variant displayed.

Explain the role PfEMP1/var genes in immune evasion

• Because PfEMP1 varies, the host's immune system struggles to recognize and clear infected RBCs.

• As soon as antibodies form against one PfEMP1 type, the parasite switches var genes, presenting a new version that is not yet recognized by the immune system.
  This allows the parasite to persist in the bloodstream for weeks or months.

Explain the role PfEMP1/var genes in pathogenesis

PfEMP1 helps infected RBCs stick to blood vessel walls — a process called cytoadherence.
This prevents the infected RBCs from being filtered out by the spleen.
However, this also leads to blockage of small blood vessels, especially in organs like the brain (cerebral malaria) or placenta (placental malaria)

Explain the current thinking about how a single var gene is expressed at a time

• Var genes are expressed one at a time ( mutually exclusive expression )

Involves:

• epigenetically regulation
• nuclear positioning
• noncoding RNA

Epigenetic Regulation (Gene "Silencing" and "Activation")

• Var genes are mostly silenced through heterochromatin—a tightly packed form of DNA that prevents transcription.

• The active var gene is in euchromatin, a relaxed, transcriptionally active form.

• Specific histone modifications (e.g. methylation or acetylation of histone H3) determine whether a var gene is silenced or active.

Spatial Nuclear Localisation/ positioning

• The active var gene moves to a special transcriptionally active region of the nucleus.

• All other var genes are located in repressive heterochromatic clusters at the nuclear periphery.

• This spatial organisation helps ensure only one gene has access to transcription machinery at a time.

swithcing mech

• Occasionally, the parasite switches the active var gene, likely through changes in chromatin structure and relocation to the active nuclear zone.

• This process is not random—some var genes are more likely to be activated than others depending on the parasite stage and host immune pressure.

Explain why previously immune women in endemic malarial regions experience malarial disease during their first pregnancy

On- During pregnancy, Plasmodium falciparum expresses a special variant of the PfEMP1 protein, called VAR2CSA.

• VAR2CSA enables infected red blood cells to adhere to chondroitin sulfate A (CSA) on the placental surface, instead of the usual endothelial receptors.

• This allows the parasite to sequester in the placenta, avoiding clearance by the spleen and triggering local inflammation.

Women who have had malaria before are typically immune to common PfEMP1 variants. But they have not developed antibodies to VAR2CSA, because it is only expressed in the placenta—which they had never had before pregnancy.
So, during their first pregnancy, they are immunologically naïve to VAR2CSA, making them vulnerable to placental malaria.

Describe the process Plasmodium falciparum uses to export proteins

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1. Protein Synthesis Inside the Parasite: All exported proteins are first synthesized in the parasite's cytoplasm using its ribosomes.
   Many of these proteins contain a PEXEL motif (Plasmodium Export Element), a short amino acid sequence (RxLxE/Q/D) that signals them for export.

2. Processing by Plasmepsin V:
   Proteins with a PEXEL motif are transported into the parasite's endoplasmic reticulum (ER).
   In the ER, the enzyme Plasmepsin V cleaves the PEXEL sequence at a specific site.
   This cleavage is required for proper targeting to the export pathway.

3. Transport Through the Parasite's Secretory Pathway:
   After PEXEL processing, proteins are trafficked via the Golgi apparatus and secretory vesicles toward the parasite plasma membrane (PPM)

4. Passage Through the Parasitophorous Vacuole
   The parasite resides in a membrane-bound compartment inside the RBC called the parasitophorous vacuole (PV).
   Exported proteins must cross the parasitophorous vacuole membrane (PVM) to reach the host cytoplasm.

5. PTEX Complex Mediates Translocation:
   The Plasmodium Translocon of Exported proteins (PTEX) is a molecular machine embedded in the PVM.
   It translocates exported proteins across the PVM into the host RBC cytosol.
   PTEX is composed of several key proteins:
   HSP101: an unfoldase that threads proteins through
   EXP2: the pore-forming channel
   PTEX150 and others: structural support

6. Remodeling the Host RBC:
   Once inside the RBC, these exported proteins help:
   Form Maurer's clefts (transport hubs)
   Build knobs on the RBC surface (to present PfEMP1)
   Modify RBC rigidity and adhesion properties
   This enables the parasite to adhere to blood vessel walls and evade spleen clearance.

Explain why protein export machinery is a target for malarial drug development

1. Essential for Parasite Survival
   Exported proteins remodel the host red blood cell (RBC) to:
   Enable nutrient uptake
   Build PfEMP1 knobs for immune evasion
   Prevent clearance by the spleen
   Without protein export, the parasite cannot survive or replicate within RBCs.

2. Unique to the Parasite
   The export machinery (especially the PTEX complex and Plasmepsin V) is absent in human cells.
   This makes it a highly selective drug target—drugs that block it are less likely to harm human proteins, minimizing side effects.

3. Slows Antigenic Variation
   Blocking protein export interferes with PfEMP1 trafficking, which helps the parasite avoid the immune system.
   Preventing PfEMP1 display reduces cytoadherence and may reduce the severity of disease (e.g., cerebral or placental malaria).

List and briefly describe the three types of potential malaria vaccine

1. Pre-erythrocytic Vaccines

2. Blood-stage Vaccines

3. Transmission-blocking Vaccines

Pre-erythrocytic Vaccines

Target: Sporozoite stage and liver stage (before red blood cell infection) to prevent the parasite from infecting liver cells or developing further after mosquito bite.

RTS,S and R21 targets CSP (circumsporozoite protein) on the surface of sporozoites.

moderate efficiency that declines rapidly so requires boosters

Blood-stage Vaccines

Target: Merozoite stage (parasite inside red blood cells) to limit parasite replication in red blood cells to reduce disease severity and symptoms

Targets: Surface proteins RH5 that may not prevent infection but helps reduce parasitemia and severity.

Drawback is to do with the polymorphism of RBC surface, so look for alternate ligands

Transmission-blocking Vaccines

Target: Sexual stages of the parasite inside the mosquito to prevent the parasite from developing in the mosquito, blocking transmission to others.

Antigens like Pfs25, which are expressed in gametocytes or zygotes in mosquitoes.

Provides community-level protection, not direct benefit to the vaccinated individual.

Explain why the malarial blood stage is a good target for vaccination

The malarial blood stage is considered a good target for vaccination because it is the stage responsible for the clinical symptoms and pathology of malaria, and offers several accessible targets for immune intervention

1. The blood stage (when merozoites infect red blood cells) is responsible for fever, anemia, and severe complications like cerebral malaria.
   Targeting this stage can reduce illness, hospitalization, and death, even if it doesn't fully prevent infection.

2. People living in endemic areas often develop partial immunity to blood-stage parasites after repeated infections.
   This suggests the human immune system can recognize and control this stage, and that vaccination could mimic this immunity.

3. Surface Antigens Are Accessible
   Merozoites and infected red blood cells express proteins on their surfaces (e.g., MSP1, PfRH5, and PfEMP1).
   These proteins are exposed to antibodies, making them promising targets for vaccine-induced immune responses.

Describe the process by which merozoites invade red blood cells

1. Initial Contact
   The merozoite (free blood-stage parasite) makes a loose, reversible contact with the red blood cell.
   This is mediated by merozoite surface proteins (MSPs), which help the parasite sense and attach to the RBC membrane.

2. Reorientation
   The merozoite reorients so that its apical end (containing invasion organelles like rhoptries and micronemes) faces the RBC surface.
   This step is crucial to initiate tight, directed entry.

3. Formation of Tight Junction
   Specialized proteins are secreted from micronemes and rhoptries, including:
   EBA (erythrocyte-binding antigens)
   PfRH (reticulocyte-binding homologues)
   These bind to specific receptors on the RBC surface (e.g., glycophorins), forming a tight junction—a ring-like structure through which the parasite enters.

4. Active Invasion
   The merozoite uses its actin-myosin motor (glideosome) to pull itself into the RBC through the tight junction.
   During this process, the parasite creates a parasitophorous vacuole—a membrane-bound compartment that protects it inside the host cell.

5. Sealing and Modification
   Once inside, the RBC membrane reseals, and the parasite begins to remodel the red blood cell using exported proteins.
   This creates a hospitable environment for growth, replication, and immune evasion.

Explain how selective pressure can alter receptor-ligand expression in Plasmodium

Selective pressure can come from:
Host immune responses
Genetic diversity in human RBC receptors
Antimalarial drugs or vaccines targeting specific ligands
Environmental changes (e.g. transmission intensity)

In response, Plasmodium can:
Switch expression between different ligand families
Mutate ligand genes to evade antibodies
Preferentially use alternative invasion pathways

Describe the basis of the most common rapid diagnostic test for malaria and explain why new tests are required

The most common malaria RDT detects Plasmodium falciparum by targeting the HRP2 protein:

Target: Histidine-Rich Protein 2 (HRP2)
HRP2 is a water-soluble protein secreted by P. falciparum during its blood stage.
It is abundant in infected individuals and easily detectable in blood samples.

A small blood sample (e.g., from a finger prick) is applied to the test strip.
The strip contains antibodies specific to HRP2 conjugated to a dye.
If HRP2 is present, it binds to these antibodies and forms a visible line—similar to a pregnancy test.
A control line ensures the test is functioning correctly.

new ones are required as

1. Some P. falciparum strains have deleted the hrp2 (and/or hrp3) gene.
   These strains do not produce HRP2, causing false-negative results.

2. HRP2 tests only detect P. falciparum.
   They miss infections caused by:
   P. vivax

3. Antigen Persistence
   HRP2 can remain in the blood for weeks after successful treatment.
   This leads to false positives,

4. Sensitivity and Performance
   Some tests are less reliable at low parasite densities, especially in early infection.
   Temperature and humidity in field conditions may affect test accuracy.