The burden of malaria
Current diagnostic technologies and the challenges of detecting the ‘last’ parasite
LoD (p/µL or ng mL−1) | Sensitivity (%) (95% CI) | Specificity (%) (95% CI) | Cost ($US/test) | Time | Other requirements | ||
---|---|---|---|---|---|---|---|
Instrument | Test | ||||||
Case management | |||||||
Microscopy | Expert: 4–20 [18] | Depends on microscopist | ~ 3000 | 0.12–0.40 [19] | 60 min [18] | Trained personnel, microscope, Giemsa stain [18] | |
Average: 50–200 [19] | |||||||
RDTs | Existing RDTs: 100 p/µL [22] Latest product: 80 pg/mL for PfHRP2 [21] | > 85% depending on species [19] | > 99% [19] | No need for expensive instrument | 0.55–1.50 [18] | 20 min [20] | Test kit, appropriate storage conditions [18] |
Surveillance | |||||||
RDTs | Latest product: 80 pg/mL for PfHRP2 [21] | > 85% depending on species [19] | > 99% [19] | No need for expensive instrument | 0.55–1.50 [18] | 20 min [20] | Test kit, appropriate storage conditions [18] |
PCR | 26 (real-time) [10] | 100% [23] | > 99% [10] | Real-time instrument > 20,000 [25] | 1.5–4.0 [24] | Standard > 6 h | Thermocycler, cold chain, power, reagent grade, water |
− 0.5 to 5. 0 [24] | |||||||
LAMP | 47 (real-time) [10] | 83.3% [22] | > 99% [22] | Conventional PCR and LAMP ~ 5000 [25] | 0.40–0.70 [24] | 60 min | Heat source for amplification and DNA extraction |
≥ 1 [23] | 97.3% [24] | > 85% [23] |
Microscopy
Rapid diagnostic tests
Advantage | Disadvantages |
---|---|
Easy to use | Deletion of the Pfhrp2 gene leads to false negative RDTs (particularly in populations in the Amazon region) |
Low cost | Lack of adequate sensitivity for detection of infection in asymptomatic individuals and/or prozone effect |
Quick result delivery time (< 20 min) | Lack of heat stability when being stored in endemic settings |
Portable and disposable | Inability to differentiate non-Pf malaria |
Require minimal laboratory infrastructure, power or external equipment | Inability to distinguish current and past infections |
Quick training | Inability to quantify parasite density, especially for assessing severity of illness or monitoring treatment efficacy |
Promising and alternative technologies for malaria detection
Technology | Product | Developer | Description | Type of detection | Performance | Turn-around time | Sample type | Environmental requirements | Cost per test | Cost per instrument | Power/labour/infrastructure requirements | Result display and storage | Quality control |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Microscopy | Parasight | Sight Diagnostics Ltd, 2014 | Automated microscopy suitable for processing of multiple malaria | Slide reading | Under way | n/a | Blood smear | n/a | n/a | n/a | n/a | ||
Malaria RDTs** | Fio-net | Fio Corporation, 2012 | Universal RDT reader and cloud information services to improve malaria RDT quality assurance and malaria surveillance | Combination of mobile diagnostics (mobile universal reader) with cloud information services | Automated and customising reports Sensitivity and specificity are functions of the RDTs being read | RDTs processing time is dependent on manufacturer’s recommendation Data upload within minutes Daily quality control needed | Depending on RDTs’ manufacturers | Subject to RDTs manufacturers’ recommendations 5–40 °C | Similar to pre-paid cellphone plans | Battery powered Basic 1 day training needed | On screen and web portal | CE marked | |
UMT | Fyodor Biotechnologies, 2015 | A senstitive and specific lateral flow assay detecting novel Plasmodium proteins shed in the urine of febril malaria patients | Dipstick technology (lateral flow assay) | LOD 125 parasites/µL | ~ 20 min | 100 µL urine | n/a | n/a | Usable by lay people | n/a | n/a | ||
Holomic Rapid Diagnostic Reader | Holomic LLC, 2013 | Universal RDT reader attachment for smartphones and software to read RDTs and transmit result to a secure cloud information service | Portable, smartphone-based lateral flow immunoassay reader | Quantitative and qualitative | RDTs processing time is dependent on manufacturer’s recommendation Data upload within seconds | Depending on RDTs’ manufacturers | Subject to RDTs manufacturers’ 5–40 °C | Customisable | $US500 | Battery powered Basic < 0.5 day training needed | User interface of the smartphones application | Class I medical device | |
Nucleic acid detection | LAMP Malaria Diagnostic Kit | Eiken Chemical Ltd and FIND, 2012 | Commercial LAMP test kit containing primers and reagents needed to run assays using benchtop laboratory equipment | Isothermal DNA amplification Fluorescence of visual detection | For pan-LAMP: 97.0% sensitivity For Pf-LAMP: 93.3% sensitivity 85.0% specificity | 60 min | 30–60 µL blood | Stable for 12 months at < 30 °C | $US5 | $US10’000 | Electricity (batter-powered possible) 4 days of training required | Turbidimeter and software | CE marked Positive and negative controls included |
illumigene LAMP | Meridian Bioscience | An automated and compact LAMP technology to qualitatively detect Plasmodium spp. DNA in human whole blood samples | Isothermal DNA amplification | Sensitivity 100% Specificity 89.3% | < 50 min | Human whole blood | Stable for 12 months at 2–30 °C | n/a | Does not require specialised laboratory equipment | n/a | CE marked | ||
MicroPCR | Tulip Group and Bigtec Labs, 2013 | POC real-time quantitative PCR instrument | Fluorescent probe-based real-time PCR | > 99% sensitivity and specificity LOD 2 parasites/µL blood | 45–60 min | 100 µL blood | 15–30 °C | $US15 | $US8000 | Battery powered 1–2 days training required | 5000 test results can be stored internally, cloud information available | CE marked | |
Truelab | Molbio, 2013 | A quantitative micro PCR platform containing all equipment and reagents needed for point-of-care applications | Using the proprietary magnetic nanoparticles to capture DNA | n/a | < 60 min | Whole blood | n/a | n/a | A customised micro printer is available | n/a |
Specifications for a new generation of malaria RDTs
Microfluidic technology for malaria POC testing
Application | Concept/detection principle | Biomarker/target | Limit of detection | Performance | Time (min) | Refs | |
---|---|---|---|---|---|---|---|
Sensitivity (%) | Specificity (%) | ||||||
Molecular analysis | Paper-based LAMP |
P. falciparum
| 5 p/µL | 61% | 98% | 45 min | [81] |
P. vivax
| 81% | 98% | |||||
P. pan
| > 80% | > 98% | |||||
Continuous flow PCR |
P. falciparum
| 2 p/µL | 97.40% | 93.80% | n/a | [86] | |
< 1 p/µL | n/a | n/a | 2.5 h | [87] | |||
Cell deformation mechanism | Inertial focusing |
P. falciparum
| 2–10 p/µL | n/a | n/a | 400 µL/min | [88] |
Inertial microfluidics | P. falciparum iRBCs | 2 cells/min | n/a | [89] | |||
Non-inertial lift effect | P. falciparum ring stage iRBCs | Enrichment factor of 4.3 | n/a | [90] | |||
Throughput 12,000 cells/h | |||||||
Electrical detection | Electrical conductivity of iRBCs is significantly higher than healthy RBCs | P. falciparum ring stage | n/a | n/a | [91] | ||
Optofluidic-flow analyser that can measure the optical absorption of RBCs in P. falciparum infected blood sample |
P. falciparum
| 1712 RBCs/s | n/a | 3 min | [92] | ||
2.96% parasite density | |||||||
Naked-eye screening of in-meso detection of hemozoin crystallites based on birefringence | Hemozoin crystals produced by P. falciparum | n/a | ~ 12 min | [58] | |||
Optical detection | Visual detection of colored assay spot on a disposable microfluidic card based on a flow-through membrane immunoassay | Malaria PfHRP2 | 10–20 ng/mL | n/a | 1–5 min | [79] | |
Paper-based catridge containing detection areas for both thin and thick smears |
P. falciparum
| 100 p/µL | n/a | 30 min | [93] | ||
Magnetic detection | Cell enrichment microfluidics combined with magnetic relaxometry detection | P. falciparum ring stage parasites | 5% parasite density | n/a | 15 min | [54] | |
Detection of hemozoin in iRBCs by magnetic resonance relaxometry | Hemozoin in iRBCs in P. falciparum infections | < 10 p/µL | n/a | Few mins | [94] |
Immunodiagnostics on microfluidic platforms for malaria detection
Sample pre-concentration
Flow control
Detection
Molecular testing on microfluidic platforms for malaria detection
Sample pre-treatment
Heating systems
Paper-based microfluidics
Interfacing microfluidic-based analysis with networked mobile devices
Optical detection | Data analysis | Signal transduction | Target biomarker | Sample | Platform | Performance | Refs. |
---|---|---|---|---|---|---|---|
Phone LED and camera + 4 external lenses and mirrors | Mie scattering simulation online | Immunoagglutination (Mie light scattering) | PfHRP malaria biomarker | Human blood | Microbeads | 1 pg/mL–10 ng/mL | [144] |
LOD 1 pg/mL | |||||||
Computational power + external optical fiber + LED | Phone application | Fluorescence | Genomic DNA | Escherichia coli and Staphylococus aureus | Microfluidics | Comparable to that of commercial PCR | [138] |
Phone camera | Phone app | Colorimetry | HE4 (ovarian cancer biomarker) | Urine | Microchip | 89.5% sensitivity, 90% specificity | [139] |
2 external LEDs + phone camera | Phone app | Colorimetry | Peanut | Cookies | Sample holder | < 1 ppm | [140] |
External LED + phone camera + additional lens | Phone application | Fluorescence |
Escherichia coli
| Milk, water | Glass capillary | 5–10 cfu/mL | [141] |
External LED and optical fibers | Phone app | Immunochromatography (Mie scatter) | Thyroid stimulating hormone | Human serum | Nitrocellulose test strip | 0.31 mIU/L | [142] |
Phone camera + external LED | Computer | Colorimetry | Human IgG | Human IgG sample | Microfluidics, silver deposition | n/a | [143] |
Snap-on attachment (lens + LEDs) + phone camera | Phone app | Immunochromatography | Malaria biomarkers | Whole blood | Rapid test diagnostic strips | 4 × dilution c.f. RDTs | [145] |
3 external attachments + lenses + LED + phone camera | Phone application | Fluorescence | Cell count | Blood | Sample holder | 600–2500 white cells/image | [146] |
400–700 red cells/image | |||||||
Phone camera | Phone app | Colorimetry | pH | Test strip | n/a | [147] | |
External LEDs and photodiode | Phone app | Colorimetry | Glucose | Urine | Paper strips | 0–250 mg/dL | [148] |
LOD 10 mg/dL | |||||||
Snap-on attachments (lens + LED) + phone camera | ImageJ on computer | Fluorescence | Prostate specific antigen (PSA) | Whole blood | Microfluidics | Dynamic range 0.08–60 ng/mL | [149] |
LOD 0.4–0.04 ng/mL |