Week 1 Lecture 1 - Malaria Flashcards
Life Cycle Stages of Malaria in Humans
Shizont
Trophozoite
Gametocyte
Malaria - Clinical
Mild-asymptomatic
Acute self-limiting febrile, myalgia, nausea, vomiting, diarrhoea
Severe, life-threatening
Poor prognosis
- severe anaemia, respiratory distress, cerebral malaria, jaundice, renal failure, shock etc.
Haematological Changes due to Malaria
Anaemia
- sequestration of parasitised RBC within bone marrow
- maturation defects
- dyserythropoiesis (P. vivax)
- destruction of parasitised RBC at merogony
- enhanced splenic clearance of infected RBC
Thrombocytopenia
- common P. vivax, P. falciparum
- decreased platelet survival (2-4 days)
- enhanced splenic clearance, destruction/sequestration
Leukopenia
- mild in uncomplicated cases
Leukocytosis
- associated with severe P. falciparum malaria
Laboratory Diagnosis of Malaria
Thrombocytopenia (<120 x 10^9/L)
Leukopenia (<4.0 x 10^9/L)
Demonstration of parasites in blood film:
- thin film (x3 for screening)
- thick film (x2 for conformation)
Thick Film Preparation
Place a drop of blood onto microscope slide & spread the drop until it is about 1-2 cm2
Adjust thickness so that its just possible to see through it
Allow the films to dry
DO NOT fix thick films
Stain:
- prepare a 10% solution of Giemsa stain in phosphate buffer pH 7.2
- filter into a coplin jar
- stain slides for 20 min
- allow to air dry
Thin Film Preparation
Thin films are made in the standard manner:
- air dry
- fix
- stain with May-Grünwald & Giemsa (or equivalent
Romanowsky stain)
pH of the stain:
- slightly alkaline stain is recommended (pH 7.2)
- staining in an acid (pH < 6.8) may fail to show parasites
Thick Film vs Thin Film
Thick
- more likely to detect Plasmodium organisms
- organisms distorted so more difficult to determine species
Thin
- allows better assessment of morphology of parasites
- allows speciation
- low density infections may be difficult to detect organisms
Parasitaemia
No. parasites 100-10^6 per uL of blood
Combined circulating RBC & sequestered RBC
Parasitaemia determined as no. parasitised RBC per 1000 RBC
Can be used to follow course of disease/response to treatment
4 Most Common Species of Malaria
Plasmodium falciparum
Plasmodium vivax
Plasmodium ovale
Plasmodium malariae
In Australia: P. vivax > P. falciparum > P. ovale and P.malariae
Malaria Species Characteristics - RBC Enlargement
Plasmodium vivax - yes
Plasmodium falciparum - none
Plasmodium ovale - yes
Plasmodium malariae - none
Malaria Species Characteristics - Trophozoite
Plasmodium vivax
- size > 1/3 of RBC
- multiple parasites rare
- rough shape; single chromatin dot
Plasmodium falciparum
- size <1/3 of RBC
- multiple parasites common
- delicate shape; often double chromatin dot
Plasmodium ovale
- size >1/3 of RBC
- multiple parasites rare
- rough shape; single chromatin dot
Plasmodium malariae
- size >1/3 of RBC
- multiple parasite rare
- compact shape; inverted chromatin dot
Malaria Species Characteristics - Schizont
Plasmodium vivax
- frequency common
- random configuration
- no. of merozoites is 12-24
Plasmodium falciparum
- frequency very rare
- random configuration
- no. of merozoites is 8-24
Plasmodium ovale and Plasmodium malariae
- frequency common
- daisy-head configuration
- no. of merozoites is 8-12
Schuffner’s Dots
Visible in Romanowsky-stained blood smears as multiple brick-red dots
Presence of Schuffner’s Dots in Malaria species:
- Plasmodium vivax: yes, fine
- Plasmodium falciparum: no
- Plasmodium ovale: yes, large
- Plasmodium malariae: no
Plasmodium vivax
Most commonly seen Plasmodium species in Australia
Usually not life threatening
Invades reticulocytes
~48 hour cycle
Diagnostic points:
- infected red cells are typically enlarged
- ring forms usually large ~ 1/3 size of red cell
Usually responds well to treatment
Plasmodium falciparum
Invades reticulocytes & mature RBCs
~48 hour cycle
Typically greater parasitaemia than other Plasmodium species
Most pathogenic
- cerebral sequestration
- sequestration other organs
- life threatening => multiple organ failure, death
- children most vulnerable
Diagnostic points
- RBC typically not enlarged
- small, fine ring forms
- occasional gametocytes recognised
Treatment often difficult due to drug resistance
Plasmodium Ovale
RBC enlarged
~48 hour cycle
Typically not life-threatening
Diagnostic Points
- red cells enlarged
- comet forms common (top right)
- rings large and coarse
- Schuffner’s dots, when present, may be prominent
- mature schizonts similar to those of P. malariae but larger and more coarse
Plasmodium Malariae
RBC not enlarged
~72 hour cycle
Typically not life-threatening
Diagnostic Points
- ring forms may have a squarish appearance
- band forms are a characteristic of this species
- red cells are not enlarged
- chromatin dot may be on the inner surface of the ring
Plasmodium knowlesi
Definitive host: macaque monkeys => Zoonotic
Present throughout SE Asia
Causes severe disease in humans
Lifecycle of 24h
Infects young & mature RBC
Risk Factors for Misidentification of Malaria
Poorly prepared/stained preparations
Deteriorated organisms
Low number of parasites
Mixed infections
Novel organisms
Rapid Diagnostic Tests
RDTs use small blood samples obtained by finger prick or by venepuncture
In general, a blood specimen to be tested (2–50μl) is lysed in buffer solution containing one or more malaria specific ‘detection antibodies’.
Important to follow up with thick and thin films on negatives especially with suggestive history/clinical signs/FBC results
PCR Test
Useful to detect low concentration of parasites
Possible to detect <10 parasites per 10uL of blood
Used as an adjunct to the examination of blood films for the diagnosis & speciation of malaria
Useful for detection of:
- multiple infections
- species with similar morphology
- residual infection especially during/post-treatment
PCR Results for each Species of Malaria
P. vivax - 120bp
P. falciparum - 205bp
P. ovale - 800bp
P. malariae - 144bp
