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" Development and Evaluation of Diagnostics for Malaria Control "
Unwin, Vera Tuasina
Adams, Emily
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Latin Dissertation
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| Language of Document
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English
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| Record Number
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1112121
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TLpq2518399741
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| Main Entry
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Adams, Emily
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Unwin, Vera Tuasina
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| Title & Author
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Development and Evaluation of Diagnostics for Malaria Control\ Unwin, Vera TuasinaAdams, Emily
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| College
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The University of Liverpool (United Kingdom)
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| Date
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2020
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| student score
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2020
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| Degree
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Ph.D.
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| Page No
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267
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| Abstract
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Introduction WHO malaria elimination targets require sustained and robust surveillance systems to identify difficult-to-diagnose, low-density malaria infections which threaten control efforts. The WHO 'T3: Test, Treat and Track' initiative further reinforces the importance of these principles (Cox 2010). Early detection and treatment of malaria is especially important during pregnancy given its adverse outcomes. Prior to administration of antimalarials, the WHO recommends confirmation of diagnosis of malaria. The current gold standard for this is still microscopy, with rapid diagnostic tests (RDTs) commonly employed where microscopy services are limited. However, both methods are insufficiently sensitive to detect low-density or "sub-microscopic" infections. Individuals harbouring these infections act as an infectious transmission reservoir contributing to ongoing transmission in low-transmission/near-elimination settings. Unfortunately, only molecular methods such as qPCR- which are largely restricted to use in well-supported laboratories- are currently suited to diagnosing these individuals. Aims There is an urgent need for sensitive diagnostic tools for malaria that are better suited to lowresource settings. The overall objective of the thesis is to investigate new diagnostic tests, molecular techniques, and adaptations to enhance the detection of low-density malaria infections in limited-resource settings. This thesis aims to explore new diagnostics suitable for 1) molecular surveillance and 2) improved point-of-care (POC) diagnosis. The specific objectives are to: 1) Assess the suitability of high-throughput molecular assays for differentiating Plasmodium species (Chapter 2) 2) Develop a new molecular test for malaria control applications on a portable, field-friendly platform (Chapter 3) 3) Characterise the diagnostic performance of a new RDT for detecting low density malaria during pregnancy in low and high transmission settings (Chapter 4-6) Methods and key findings The differentiation of the main species of Plasmodium is important not only for the treatment of liver stages unique to P. vivax, but also for accurate surveillance of malaria epidemiology. Chapter 2 explores molecular techniques; namely, High Resolution Melt Curve Analysis (HRMCA) and real-time qPCR, to develop a multiplex qPCR suitable for the detection of the species of malaria most prevalent in the study areas of interest: Indonesia and Kenya. As a result, a new 4-plex qPCR assay was optimised and utilised in a large clinical trial to confirm infection amongst asymptomatic women in Indonesia. The feasibility of using whole blood directly in the 4-plex was also demonstrated, as was the transfer of the qPCR to the more portable Magnetic Induction Cycler (MIC™) device (chapter 3). Using the multiplex qPCR as a reference standard, the diagnostic performance of a new ultrasensitive P. falciparum RDT (uRDT) was retrospectively assessed in a Stop Malaria in Pregnancy clinical trial in Indonesia (chapter 4). The uRDT's performance was compared to that of the widely used Pf/Pan CareStart RDT (csRDT). The overall similar performance of the two RDTs (uRDT 20% sensitivity, 98% specificity and csRDT 23%, 96%) led to an investigation into antigenic mutations that may account for RDT insensitivity (chapter 5). The results from this study suggest a high proportion of infections that were qPCR positive and RDT negative are attributable to HRP2 deletions. The uRDT performance in comparison to csRDT was further investigated amongst pregnant women in Kenya, to investigate the effects of transmission intensity. The sensitivity of the uRDT (79.9%) was significantly higher than that of the csRDT (74%), although the specificity was significantly lower (uRDT; 90.4% and csRDT; 93.0%). Overall, the performance of the two tests (as indicated by the Diagnostics Odds Ratios, Kappa values and AUROC values) were nonsignificantly different to each other. Finally, a novel assay for mosquito insecticide resistance was developed on the portable pointof-care Genedrive® platform, demonstrating its potential for use as a field-friendly diagnostic tool for malaria control. Conclusions The uRDT evaluations performed here contribute to critical understanding of how these tests perform in pregnant women in different transmission settings. The results suggest that in low transmission settings this test may not offer an improvement on currently used RDTs, particularly for screening asymptomatic infections during pregnancy. In contrast, the uRDT may be better suited for this role in higher transmission settings, such as in Kenya. Although the uRDT demonstrated lower specificity in the latter scenario, the negative outcomes associated with malaria during pregnancy together with the safety of current antimalarial therapies mitigate the risks of unnecessary treatment. The poor sensitivity of both HRP2-targetting RDTs do not support their use for screening for malaria during pregnancy, with the caveat that the off-label use of stored samples may have affected their sensitivity. The evidence of HRP2 deletions is the first report of these mutations in Indonesia. Although further confirmatory studies are needed to strengthen these findings, they flag a potential barrier to the current test and treat policies for screening pregnant women in Indonesia. The work highlights the urgent need for wider and more in-depth HRP2 studies in this region to better inform these programs. Overall, this thesis tackles several pertinent challenges around improving malaria diagnostics. It highlights issues that need to be considered during the product design, development and assessment phases. It also incorporates useful evaluation studies of novel tests with the potential to make malaria diagnostics more accessible. Its translational approach is suited to product developers, as well as those influencing control/ surveillance programs and associated policy.
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Erythrocytes
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Infections
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Malaria
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Polymerase chain reaction
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Pregnancy
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Vaccines
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