Malaria PCR Detection in Cambodian Low-Transmission Settings: Dried Blood Spots Versus Venous Blood Samples

Abstract

Abstract. In the context of malaria elimination, novel strategies for detecting very low malaria parasite densities in asymptomatic individuals are needed. One of the major limitations of the malaria parasite detection methods is the volume of blood samples being analyzed. The objective of the study was to compare the diagnostic accuracy of a malaria polymerase chain reaction assay, from dried blood spots (DBS, 5 μL) and different volumes of venous blood (50 μL, 200 μL, and 1 mL). The limit of detection of the polymerase chain reaction assay, using calibrated Plasmodium falciparum blood dilutions, showed that venous blood samples (50 μL, 200 μL, 1 mL) combined with Qiagen extraction methods gave a similar threshold of 100 parasites/mL,~100-fold lower than 5 μL DBS/Instagene method. On a set of 521 field samples, collected in two different transmission areas in northern Cambodia, no significant difference in the proportion of parasite carriers, regardless of the methods used was found. The 5 μL DBS method missed 27% of the samples detected by the 1 mL venous blood method, but most of the missed parasites carriers were infected by Plasmodium vivax (84%). The remaining missed P. falciparum parasite carriers (N = 3) were only detected in high-transmission areas.BACKGROUND

Genetic diversity in twoPlasmodium vivaxprotein ligands for reticulocyte invasion

“…DNA was extracted from blood spots with Instagene Matrix reagent (BioRad, Marnes-la-Coquette, France) or from whole blood samples using the QIAamp DNA Blood Mini Kit (Qiagen, Courtaboeuf, France), according to the manufacturer's instructions. Molecular detection and identification of Plasmodium parasites were performed by using real-time PCR targeting the cytochrome b gene as described previously [47,48]. Table).…”

Detection of Malaria in Healthy Blood Donors using PCR in an Endemic Area in Yemen

“…For two decades, the sensitivity of malaria diagnostic tests has considerably been improved with the introduction of molecular assays (Canier et al, 2015). Methods based on molecular biology have been used to detect different types of Plasmodium by PCR, such as nested PCR (Batista-dos-Santos et al, 2012).…”

“…Methods based on molecular biology have been used to detect different types of Plasmodium by PCR, such as nested PCR (Batista-dos-Santos et al, 2012). The performance of nested PCR as a ma laria diagnostic tool is excellent with respect to its high accuracy, sensitivity, specificity, and ability to discriminate Plasmo dium species (Canier et al, 2015). PCR technique can detect parasites below the threshold levels of microscopy; when performed under optimal conditions, PCR can detect parasitaemia as low as 0.01-1 parasite/μL of blood (Tangpukdee et al, 2009).…”

Performance of Ultrasensitive Rapid Diagnostic Tests for Detecting Asymptomatic Plasmodium falciparum

“…Both village malaria workers and survey data collectors were provided with training on how to take blood samples and carry out RDTs correctly as well as preparing filter paper and preprepared 96-well plates for subsequent PCR analysis. Nucleic material extraction from dried blood spot and screening for any malaria species infection and then specific P. falciparum infection were carried out according to the methodology of Canier et al 11 Results of RDTs were recorded on paper-based forms by field workers, uploaded into an Excel (Microsoft Corporation, Redmond, WA) database, and then merged with the results of qPCR analysis using the unique participant identification number. The descriptive statistical analysis was carried out using Stata 15.1 (StataCorp, College Station, TX).…”