Comparison of a Genus-Specific Conventional PCR and a Species-Specific Nested-PCR for Malaria Diagnosis Using FTA Collected Samples from Kingdom of Saudi Arabia

Abstract

Molecular tools are increasingly accepted as the most sensitive and reliable techniques for malaria diagnosis and epidemiological surveys. Also, collection of finger prick blood spots onto filter papers is the most simple and affordable method for samples preservation and posterior molecular analysis, especially in rural endemic regions where malaria remains a major health problem. Two malaria molecular diagnostic tests, a Plasmodium genus-specific conventional PCR and a Plasmodium species-specific Nested PCR, were evaluated using DNA templates prepared from Whatman-FTA ®cards' dry blood spots using both, Methanol-fixation/Heat-extraction and FTA® commercial purification kit. A total of 121 blood samples were collected from six Saudi southwestern endemic districts both, as thick and thin films for routine microscopic screening and onto FTA®cards for molecular studies. Out of the 121 samples, 75 were P. falciparum positive by at least one technique. No other species of Plasmodium were detected. P. falciparum parasites were identified in 69/75 (92%) samples by microscopic screening in health care centers. P. genus-specific PCR was able to amplify P. falciparum DNA in 41/75 (55%) and 59/75 (79%) samples using Methanol-fixation/Heat-extraction and FTA ® purification kit, respectively. P. species-specific Nested PCR revealed 68/75 (91%) and 75/75 (100%) positive samples using DNA templates were isolated by Methanol-fixation/Heatextraction and FTA ® purification methods, respectively. The species-specific Nested PCR applied to Whatman-FTA ® preserved and processed blood samples represents the best alternative to classical microscopy for malaria diagnosis, particularly in epidemiological screening.

Malaria Prevalence in a Low Transmission Area, Jazan District of Southwestern Saudi Arabia

“…Interestingly, out of these eight asymptomatic cases, seven were missed by microscopy (sub-microscopic). The sub-microscopic asymptomatic malaria infections have been demonstrated a contribution to malaria transmission in Jazan [29] and elsewhere in Thailand [30]. These sub-patent infections were most prevalent in patient above 15 years, coinciding with previous report [31].…”

Comparison of Different 18S rRNA Primers with Conventional PCR Methods in Determination of Plasmodium spp. and Evaluation of Nested PCR Method

“…Currently, molecular techniques are widely used in several studies and accepted as gold standard in laboratories depending on their very high sensitivity and specificity (21). PCR assay is a suitable technique for collection of epidemiological data and is a useful diagnostic tool particularly in regions with low incidence, in cases with low parasitemia and submicroscopic infections (17).…”

“…However, sensitivity changes depending on used method and characteristics of targeted gene (23). In several studies, it was reported that all subspecies were successfully detected with 18S rRNA based primers and their sensitivity and specificity were found to be higher than microscopic examination and rapid diagnostic tests (4,11,13,14,21,24).…”