Measuring the efficacy of anti-malarial drugs in vivo: quantitative PCR measurement of parasite clearance

Malaria Journal

Research

Khalid B Beshir¹  Rachel L Hallett¹  Robin Bailey²  Alice C Eziefula²  Colin J Sutherland³  Julie Watson⁴  Spencer D Polley⁴  Stephen G Wright⁴  Peter L Chiodini⁵ 

[1] Faculty of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine (LSHTM), WC1E 7HT, London, Keppel St, UK;Faculty of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine (LSHTM), WC1E 7HT, London, Keppel St, UK;Hospital for Tropical Diseases, Mortimer Market Centre, Capper St, WC1E 6JB, London, UK;Faculty of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine (LSHTM), WC1E 7HT, London, Keppel St, UK;Hospital for Tropical Diseases, Mortimer Market Centre, Capper St, WC1E 6JB, London, UK;HPA Malaria Reference Laboratory, LSHTM, Keppel St, WC1E 7HT, London, UK;Hospital for Tropical Diseases, Mortimer Market Centre, Capper St, WC1E 6JB, London, UK;Hospital for Tropical Diseases, Mortimer Market Centre, Capper St, WC1E 6JB, London, UK;HPA Malaria Reference Laboratory, LSHTM, Keppel St, WC1E 7HT, London, UK;

关键词: Malaria;    Parasite Density;    Artesunate;    Parasite Clearance;    Parasite Clearance Time;   

DOI  :  10.1186/1475-2875-9-312

 received in 2010-07-15, accepted in 2010-11-05,  发布年份 2010

来源: Springer

【 摘 要 】

BackgroundArtemisinin-based combination therapy, currently considered the therapy of choice for uncomplicated Plasmodium falciparum malaria in endemic countries, may be under threat from newly emerging parasite resistance to the artemisinin family of drugs. Studies in Southeast Asia suggest some patients exhibit an extended parasite clearance time in the three days immediately following treatment with artesunate monotherapy. This phenotype is likely to become a more important trial endpoint in studies of anti-malarial drug efficacy, but currently requires frequent, closely spaced blood sampling in hospitalized study participants, followed by quantitation of parasite density by microscopy.MethodsA simple duplex quantitative PCR method was developed in which distinct fluorescent signals are generated from the human and parasite DNA components in each blood sample. The human amplification target in this assay is the β tubulin gene, and the parasite target is the unique methionine tRNA gene (pgmet), which exhibits perfect sequence identity in all six Plasmodium species that naturally infect humans. In a small series of malaria cases treated as hospital in-patients, the abundance of pgmet DNA was estimated relative to the human DNA target in daily peripheral blood samples, and parasite clearance times calculated.ResultsThe qPCR assay was reproducibly able to replicate parasite density estimates derived from microscopy, but provided additional data by quantification of parasite density 24 hours after the last positive blood film. Robust estimates of parasite clearance times were produced for a series of patients with clinical malaria.ConclusionsLarge studies, particularly in Africa where children represent a major proportion of treated cases, will require a simpler blood sample collection regime, and a method capable of high throughput. The duplex qPCR method tested may fulfil these criteria, and should now be evaluated in such field studies.

【 授权许可】

Unknown   
© Beshir et al; licensee BioMed Central Ltd. 2010. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.