PCR targeting Plasmodium mitochondrial genome of DNA extracted from dried blood on filter paper compared to whole blood

Gro Ea Strøm,Sabrina Moyo,Bjørn Blomberg,Maulidi Fataki,Nina Langeland

doi:10.1186/1475-2875-13-137

Abstract

BackgroundMonitoring mortality and morbidity attributable to malaria is paramount to achieve elimination of malaria. Diagnosis of malaria is challenging and PCR is a reliable method for identifying malaria with high sensitivity. However, blood specimen collection and transport can be challenging and obtaining dried blood spots (DBS) on filter paper by finger-prick may have advantages over collecting whole blood by venepuncture.MethodsDBS and whole blood were collected from febrile children admitted at the general paediatric wards at a referral hospital in Dar es Salaam, Tanzania. DNA extracted from whole blood and from DBS was tested with a genus-specific PCR targeting the mitochondrial Plasmodium genome. Positive samples by PCR of DNA from whole blood were tested with species-specific PCR targeting the 18S rRNA locus, or sequencing if species-specific PCR was negative. Rapid diagnostic test (RDT) and thin blood smear microscopy was carried out on all patients where remnant whole blood and a blood slide, respectively, were available.ResultsPositivity of PCR was 24.5 (78/319) and 11.2% (52/442) by whole blood and DBS, respectively. All samples positive on DBS were also positive on Plasmodium falciparum species-specific PCR. All RDT positive cases were also positive by DBS PCR. All but three cases with positive blood slides were also positive by DBS.ConclusionsIn this study, PCR for malaria mitochondrial DNA extracted from whole blood was more sensitive than from DBS. However, DBS are a practical alternative to whole blood and detected approximately the same number of cases as RDTs and, therefore, remain relevant for research purposes.

Highlights

Monitoring mortality and morbidity attributable to malaria is paramount to achieve elimination of malaria
With a goal of malaria elimination becoming more relevant [1], monitoring malaria epidemiology, including submicroscopic quantities of malaria detected only by Polymerase chain reaction (PCR) [11,12], is necessary. It has been shown in some studies that PCR on DNA extracted from filter paper is less sensitive than PCR done on DNA extracted from whole blood [13], but the discrepancies found are not very remarkable
One case was positive on PCR of dried blood spots (DBS) that was negative by Rapid diagnostic test (RDT)

Summary

Introduction

Monitoring mortality and morbidity attributable to malaria is paramount to achieve elimination of malaria. Malaria killed approximately 627,000 people, mostly children in sub-Saharan Africa, in 2012 and remains an endemic disease in 97 countries It is a major health concern, especially in low-resource settings [1]. PCR has been performed on DNA epidemiology, including submicroscopic quantities of malaria detected only by PCR [11,12], is necessary It has been shown in some studies that PCR on DNA extracted from filter paper is less sensitive than PCR done on DNA extracted from whole blood [13], but the discrepancies found are not very remarkable. Ataei et al found 42.7% positive for Plasmodium species by PCR on filter paper and 53.3% positive on whole blood [7], while a study by Proux et al showed a smaller reduction in malaria detection by DBS as compared to whole blood [14]. Another study showed a ten- to 100-fold decrease in sensitivity when using DBS as compared to whole blood [15]

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