A reproducible method for extraction of Plasmodium falciparum DNA by microwave irradiation and its potential for rapid molecular diagnosis. | LitMetric
Department of Parasitology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.
Published: December 2015
AI Article Synopsis
Article Abstract
Malaria remains one of the most important communicable diseases. A rapid, simple and accurate method is a crucial part of malaria diagnosis. The aim of this study was to reevaluate the microwave irradiation method to extract DNA from Plasmodium falciparum and compare with six other existing DNA extraction methods such as QIAamp DNA mini kit (Qiagen), FTA elute card, phenol-chloroform, Chelex, Chelex without proteinase-K and Rapid boiling. Two different P. falciparum isolates were used: (i) Laboratory strains with 0.3% parasitemia and (ii) clinical isolate with 0.6% parasitemia. Each DNA extraction method was validated for the presence of P. falciparum by a routine nested and real time PCR. In order to evaluate the sensitivity of the DNA extraction by microwave, double serial dilution of P. falciparum from in vitro culture at parasitemia that ranged from 0.0001 to 0.17% were used to extract the DNA by microwave and the P. falciparum DNA was then detected by nested and real-time PCR. The nested and real-time PCR were able to detect. P. falciparum DNA at the parasitemia level as low as 0.0003% and 0.0001%, respectively. Our results can reproduce the results from earlier studies and reveal microwave as a rapid and simple tool to extract P. falciparum DNA and subsequent molecular diagnosis of malaria.
Background: Molecular methods play an important role in clinical trials assessing anti-malarial drugs and vaccines, as well as in epidemiological studies aimed at detecting Plasmodium species, especially when dealing with large sample sizes. Molecular techniques are more sensitive and generally have a higher throughput compared to the gold standard microscopy. Further optimization can be achieved with automation of nucleic acid isolation, allowing for rapid and precise extraction.
Objectives: The number of mosquito bites a person receives determines the risk of acquiring malaria and the likelihood of transmitting infections to mosquitoes. We assessed heterogeneity in biting and associated factors in two settings in Uganda with different endemicity.
Methods: parasites in blood-fed indoor caught mosquitoes were quantified using qPCR targeting the Pf18S rRNA gene.
Malaria rapid diagnostic tests (RDTs) targeting the Plasmodium falciparum histidine-rich protein 2 (PfHRP2) are widely used to diagnose P. falciparum infection. However, reports of P.
Rapid diagnostic tests (RDTs) are crucial for diagnosing malaria in resource-limited settings. These tests, which detect the histidine-rich protein 2 (PfHRP2) and its structural homologue PfHRP3, are specifically designed to identify Plasmodium falciparum. Deletion of the Pfhrp2 gene in parasite has been reported in India and other malaria-endemic countries.
* Researchers identified unique DNA sequences and protein diversity in samples with intact pfhrp2 genes, pointing to distinct amino acid repeat motifs compared to standard samples.
* The effectiveness of mass screening and treatment interventions, alongside additional measures like bed net distribution, was shown to significantly lower the risk of infection for both pfhrp2 deleted and intact P. falciparum parasites, emphasizing the need for new diagnostic methods for malaria elimination efforts by 2030.
