Pre-amplification methods for tracking low-grade Plasmodium falciparum populations during scaled-up interventions in Southern Zambia

Malaria Journal

Methodology

Rachel Daniels¹  Dyann F Wirth¹  Sarah K Volkman²  Alan Scott³  Philip Thuma⁴  Sungano Mharakurwa⁵ 

[1] Harvard School of Public Health, Boston, MA, USA;The Broad Institute, Cambridge, MA, USA;Harvard School of Public Health, Boston, MA, USA;The Broad Institute, Cambridge, MA, USA;Simmons College, Boston, MA, USA;Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA;Malaria Institute at Macha, Choma, Zambia;Malaria Institute at Macha, Choma, Zambia;Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA;

关键词: Plasmodium falciparum;    Genotyping;    Pre-amplification;    Asymptomatic;    Rapid diagnostic test;    Diagnostics;    Southern Africa;    Zambia;   

DOI  :  10.1186/1475-2875-13-89

 received in 2013-12-23, accepted in 2014-03-03,  发布年份 2014

来源: Springer

【 摘 要 】

BackgroundMalaria is receding in many endemic countries with intervention scale -up against the disease. However, this resilient scourge may persist in low-grade submicroscopic infections among semi-immune members of the population, and be poised for possible resurgence, creating challenges for detection and assessment of intervention impact. Parasite genotyping methods, such as the molecular barcode, can identify specific malaria parasite types being transmitted and allow tracking and evaluation of parasite population structure changes as interventions are applied. This current study demonstrates application of pre-amplification methods for successful detection and genotyping of residual Plasmodium falciparum infections during a dramatic malarial decline.MethodsThe study was a prospective cross-sectional design and based on a 2,000 sq km vicinity of Macha Mission Hospital in southern Zambia. Willing and predominantly asymptomatic residents of all ages were screened for malaria by microscopy during the 2005 and 2008 transmission seasons, with simultaneous collection of dried blood spots (DBS) on filter paper, and extraction of Plasmodium falciparum DNA was performed. Plasmodium falciparum infections were genotyped using a 24 SNP-based molecular barcode assay using real-time PCR. Submicroscopic parasitaemia samples were subjected to pre-amplification using TaqMan PreAmp Master Mix following the manufacturer’s instructions before SNP barcode analysis.ResultsThere was a dramatic decline of malaria between 2005 and 2008, and the geometric mean parasite density (95% CI) fell from 704/μL (390–1,271) in 2005 to 39/μL (23–68) in 2008, culminating in a large proportion of submicroscopic infections of which 90% failed to yield ample DNA for standard molecular characterization among 2008 samples. Pre-amplification enabled successful detection and genotyping of 74% of these low-grade reservoir infections, overall, compared to 54% that were detectable before pre-amplification (p <0.0005, n = 84). Furthermore, nine samples negative for parasites by microscopy and standard quantitative PCR amplification were positive after pre-amplification.ConclusionsPre-amplification allows analysis of an otherwise undetectable parasite population and may be instrumental for parasites identification, tracking and assessing the impact of interventions on parasite populations during malaria control and elimination programmes when parasitaemia is expected to decline to submicroscopic levels.

【 授权许可】

Unknown   
© Mharakurwa et al.; licensee BioMed Central Ltd. 2014. 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 credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.