Method | Key characteristics | Parasite species detectable | Advantages | Disadvantages | Reference | |
---|---|---|---|---|---|---|
Microscopic diagnosis | Conventional bright field microscopic examination | Giemsa-stained thick blood film to determine the presence or absence of malaria parasite | Plasmodium genus-specific | - Small amount of sample (blood) is required; - Able to quantify parasitaemia; - Provides prognostic information that serves as indicator for disease severity (morphological characteristics of the parasites, the maturity of asexual stages of the parasite); - Cost effective compared to molecular techniques | - Difficulties in detecting parasites in low parasite density samples (50–100 parasites/μl); - Malaria microscopist expert/well-trained personnel is needed to interpret the result (high morphological similarities between P. falciparum, P. malariae and P. knowlesi could lead to misdiagnosis and treatment delay) | [5, 13, 14, 33,34,35,36,37,38,39,40,41,42,43,44,45,46,47,48] |
Giemsa-stained thin blood film to identify the Plasmodium species | All human Plasmodium species | |||||
Quantitative buffy coat method (QBC) | Detection of malaria parasites in centrifuged peripheral blood by staining the parasite DNA with acridine orange and examination under fluorescence microscope | Plasmodium genus-specific | - Higher sensitivity (5 parasites/μl) compared to bright field microscopic examination; - Fast and easy to be performed; - Interpretation of the result is simple and requires less-trained personnel | - Difficulties in parasite species differentiation and subjective parasite quantification; - Specific equipment (fluorescence microscope) is required | ||
Partec Rapid Malaria Test (PT) | Detection of malaria parasites using test slide that is readily labelled with 4′-6-diamidino-2- phenylindole (DAPI) which binds to intraerythrocytic Plasmodium DNA, resulting in fluorescence under Partec CyScope® (fluorescence microscope) | Plasmodium genus-specific | - Easy and rapid, less labour-intensive and requires less training time for laboratory personnel; - Could be used in the field without electricity supply; - Small amount of sample (few μl) is required | - Difficulties in species differentiation; - False positive results due to the presence of non-specific artefacts or nuclei-containing cells (reticulocytes, leukocytes and bacterial cells); -Specific equipment (Partec CyScope®) is required | ||
Rapid diagnostic test (RDT) | OptiMAL® | Detection of malaria via the pLDH antigen | P. falciparum and P. vivax | Ease of use, rapid diagnosis and result interpretation, sensitive, field-deployable | Less sensitive compared to molecular diagnostic methods, heat sensitive, reduced sensitivity for non-falciparum malaria, false-negative results due to low-level expression or deletion of target antigen genes (pfhrp2) | [83] |
ParaSight-F test | Detection of malaria via the HRP-2 antigen | P.falciparum | [84] | |||
Immunochromatographic test (ICT) Malaria PF test | Detection of malaria via the HRP-2 antigen | P.falciparum | [84] | |||
SD Bioline Malaria AG Pf/Pan | Detection of malaria via the HRP-2 and pLDH antigen | P. falciparum (HRP-2), pan-malarial (pLDH) | [85] | |||
CareStartâ„¢ Malaria | Detection of malaria via the HRP-2 and pLDH antigen | P. falciparum (HRP-2), pan-malarial (pLDH) | High specificity and PPV | Low sensitivity at low parasite densities | [87] | |
Malaria pf Rapid device | Detection of malaria via the HRP-2 antigen | P. falciparum | Sensitivity and specificity comparable to those for light microscopy | [88] | ||
Ultra sensitive RDT (uRDT) | Detects HRP-2 antigen of P. falciparum malaria | P. falciparum | Higher sensitivity, specificity and ability to detect new infections faster than conventional RDT | Similar to conventional RDTs, is less sensitive compared to molecular diagnostic methods | [89] | |
Molecular diagnostic methods | Nested PCR | Targeting 18S rRNA gene | Plasmodium genus-specific | Elevated sensitivity compared to RDTs and microscopy | Cumbersome, expensive, and requires well-trained staff with stringent laboratory cleanliness to minimize risk of contamination | [101] |
Targeting 18S rRNA gene | Plasmodium genus-specific followed by nested species-specific PCR | More sensitive than microscopic examination for identification of asymptomatic malaria | [108] | |||
Targeting cytochrome b gene | Plasmodium genus-specific | Detection limit of 10 parasites/μl, better than single-round PCR and real-time methods | ||||
Semi-nested PCR | Targeting 18S rRNA gene | P. falciparum and P. vivax | More sensitive than microscopic examination for identification of sub-microscopic infections | [103] | ||
Quantitative nucleic acid sequence-based amplification (QT-NASBA) | Targeting 18S rRNA gene. Quantification was achieved by co-amplification of the RNA in the sample with one modified in vitro RNA as a competitor | P. falciparum | - Fast, sensitive, reliable, and quantitative; - Allowed for the sub-microscopic quantification; - Detection limit of 10 parasites/μl | |||
Multiplex PCR | Targeting 18S rRNA gene | P. falciparum and P. vivax | - Detection limit of 0.1 parasites/μl; - No cross-reaction between Plasmodium spp.; - Able to detect the mixed infection | [106] | ||
Real-time quantitative PCR (qPCR) | Targeting plasmepsin 4 in P. falciparum and the aspartic protease PM4 in P. vivax | P. falciparum and P. vivax | - Quantification of parasite densities; - More sensitive than microscopic examination; - Detection limit of 5.6 copies/μl; - Able to detect and quantify infections that have very low infection (0.001%) | [103] | ||
qPCR | Targeting telomere-associated repetitive element 2 and the var. acidic terminal sequence | P. falciparum | - Detection limit of 0.03 to 0.15 parasites/μl; - 10× more sensitive than standard 18S rRNA qPCR | [110] | ||
Reverse transcription-polymerase chain reaction (qRT-PCR) | Targeting 18S rRNA | P. falciparum and P. vivax | Able to detect and differentiate submicroscopic malaria infections as low as 10 parasites/ml and 18 copies/μl | [111] |